Measuring method and instrument utilizing total reflection attenuation

ABSTRACT

A plurality of measuring units each comprising a dielectric block, a metal film layer which is formed on a surface of the dielectric block and a sample holder are supported on a support. The support is moved by a support drive means to bring in sequence the measuring units to a measuring portion comprising an optical system which projects a light beam emitted from a light source, and a photodetector which detects attenuation in total internal reflection by detecting the intensity of the light beam which is reflected in total internal reflection at the interface between the dielectric block and the metal film layer. In this measuring apparatus, lots of samples can be measured in a short time.

TECHNICAL FILED

[0001] This invention relates to a measuring apparatus using attenuationin total reflection such as a surface plasmon resonance sensor forquantitatively analyzing a material in a sample utilizing generation ofsurface plasmon.

[0002] This invention particularly relates to a measuring apparatususing attenuation in total reflection which can carries out measurementon lots of samples in a short time.

[0003] This invention also relates to a measuring method employed insuch a measuring apparatus using attenuation in total reflection.

BACKGROUND ART

[0004] In metal, free electrons vibrate in a group to generatecompression waves called plasma waves. The compression waves generatedin a metal surface are quantized into surface plasmon.

[0005] There have been proposed various surface plasmon resonancesensors for quantitatively analyzing a material in a sample utilizing aphenomenon that such surface plasmon is excited by light waves. Amongthose, one employing a system called “Kretschmann configuration” is bestknown. See, for instance, Japanese Unexamined Patent Publication No.6(1994)-167443.

[0006] The surface plasmon resonance sensor using the Kretschmannconfiguration basically comprises a dielectric block shaped, forinstance, like a prism, a metal film which is formed on one surface ofthe dielectric block and is brought into contact with a sample, a lightsource emitting a light beam, an optical system which causes the lightbeam to enter the dielectric block so that the total internal reflectioncondition is satisfied at the interface of the dielectric block and themetal film and various angles of incidence of the light beam to theinterface of the dielectric block and the metal film including an angleof incidence at which surface plasmon is generated can be obtained, anda photodetector which detects the intensity of the light beam reflectedin total internal reflection at the interface and detects the state ofsurface plasmon resonance.

[0007] Various angles of incidence of the light beam to the interfacecan be obtained in the following two ways.

[0008] (1) A relatively thin light beam is deflected to impinge upon theinterface at various angles.

[0009] (2) A relatively thick light beam is caused to impinge upon theinterface in the form of convergent light so that components of thelight beam impinge upon the interface at various angles.

[0010] In the former case, the light beam which is reflected from theinterface at an angle which varies as the incident light beam isdeflected may be detected by a photodetector which is moved insynchronization with deflection of the incident light beam or by an areasensor extending in the direction in which reflected light beam is movedas a result of deflection. In the latter case, an area sensor whichextends in a direction in which all the components of light reflectedfrom the interface at various angles can be detected may be used. InJapanese Unexamined Patent Publication No. 1(1989)-138443, there isdisclosed an apparatus using the latter way in obtaining various anglesof incidence of the light beam to the interface.

[0011] In such a surface plasmon resonance sensor, when a light beamimpinges upon the interface at a particular angle of incidence θsp notsmaller than the angle of total internal reflection, evanescent waveshaving an electric field distribution in the sample in contact with themetal film are generated and surface plasmon is excited in the interfacebetween the metal film and the sample. When the wave vector of theevanescent waves is equal to the wave number of the surface plasmon andwave number matching is established, the evanescent waves and thesurface plasmon resonate and light energy is transferred to the surfaceplasmon, whereby the intensity of light reflected in total internalreflection at the interface of the dielectric block and the metal filmsharply drops. The sharp intensity drop is generally detected as a darkline by the photodetector.

[0012] The aforesaid resonance occurs only when the incident light beamis p-polarized. Accordingly, it is necessary to set the light beam toimpinge upon the interface in the form of p-polarized light.

[0013] When the wave number of the surface plasmon can be known from theangle of incidence θsp at which the phenomenon of attenuation in totalinternal reflection (ATR) takes place, the dielectric constant of thesample can be obtained. That is,${K_{sp}(\omega)} = {\frac{\omega}{c}\sqrt{\frac{{ɛ_{m}(\omega)}ɛ_{s}}{{ɛ_{m}(\omega)} + ɛ_{s}}}}$

[0014] wherein K_(sp) represents the wave number of the surface plasmon,ω represents the angular frequency of the surface plasmon, c representsthe speed of light in a vacuum, and ε_(m) and ε_(s) respectivelyrepresent the dielectric constants of the metal and the sample.

[0015] When the dielectric constant ε_(s) of the sample is known, theconcentration of a specific material in the sample can be determined onthe basis of a predetermined calibration curve or the like. Accordingly,a specific component in the sample can be quantitatively analyzed bydetecting the angle of incidence θsp at which the intensity of lightreflected in total internal reflection from the interface of the prismand the metal film sharply drops.

[0016] As a similar apparatus utilizing the phenomenon of attenuation intotal internal reflection (ATR), there has been known a leaky modesensor described in, for instance, “Spectrum Researches” Vol.47, No.1(1998), pp21 to 23 & pp26 and 27. The leaky mode sensor basicallycomprises a dielectric block shaped, for instance, like a prism, a cladlayer which is formed on one face of the dielectric block, an opticalwaveguide layer which is formed on the clad layer and is brought intocontact with a sample, a light source emitting a light beam, an opticalsystem which causes the light beam to enter the dielectric block atvarious angles of incidence so that total internal reflection conditionsare satisfied at the interface of the dielectric block and the cladlayer and various angles of incidence of the light beam to the interfaceof the dielectric block and the clad layer including an angle ofincidence at which attenuation in total internal reflection is caused byexcitation of an optical waveguide mode at the optical waveguide layercan be obtained, and a photodetector means which detects the intensityof the light beam reflected in total internal reflection at theinterface thereby detecting an excited state of the waveguide mode,i.e., attenuation in total internal reflection.

[0017] In the leaky mode sensor with this arrangement, when the lightbeam is caused to impinge upon the clad layer through the dielectricblock at an angle not smaller than an angle of total internalreflection, only light having a particular wave number and impingingupon the optical waveguide layer at a particular angle of incidencecomes to propagate through the optical waveguide layer in a waveguidemode after passing through the clad layer. When the waveguide mode isthus excited, almost all the incident light is taken in the opticalwaveguide layer and accordingly, the intensity of light reflected intotal internal reflection at the interface of the dielectric block andthe clad layer sharply drops. That is, attenuation in total internalreflection occurs. Since the wave number of light to be propagatedthrough the optical waveguide layer in a waveguide mode depends upon therefractive index of the sample on the optical waveguide layer, therefractive index and/or the properties of the sample related to therefractive index can be detected on the basis of the angle of incidenceat which the attenuation in total internal reflection occurs.

[0018] Also in the leaky mode sensor, various angles of incidence of thelight beam to the interface can be obtained in the aforesaid two ways.

[0019] The surface plasmon resonance sensor and the leaky mode sensorare sometimes used in random screening for finding a specific materialcombined with a predetermined sensing material in the field of medicinecreation. In this case, a sensing material is fixed on the film layer(the metal film layer in the case of the surface plasmon resonancesensor and the clad layer and the optical waveguide layer in the case ofthe leaky mode sensor), and a sample liquid containing a material to beanalyzed is spotted on the sensing material. Then the angle of incidenceθsp at which attenuation in total internal reflection takes place isrepeatedly measured each time a predetermined time lapses.

[0020] When the sample material (the material to be analyzed in thesample liquid) is combined with the sensing material, the refractiveindex changes with time due to combination with the sample material.Accordingly, by measuring the angle of incidence θsp at whichattenuation in total internal reflection takes place for everypredetermined time, thereby detecting states of combination of thesample material with the sensing material, whether the sample materialis a specific material to be combined with the sensing material can beknown. As combinations of such a specific material and a sensingmaterial, there has been known an antigen and an antibody. For example,there has been known measurement of detecting combination of a samplematerial with rabbit antihuman IgG antibody (sensing material).

[0021] In order to detect the state of combination of the samplematerial with the sensing material, the total reflection attenuationangle θsp (the angle of incidence θsp at which attenuation in totalinternal reflection takes place) itself need not necessarily bedetected. For example, change in the total reflection attenuation angleθsp after the sample liquid is spotted onto the sensing material ismeasured and the state of combination of the sample material with thesensing material may be measured on the basis of the change of the totalreflection attenuation angle θsp.

[0022] In the apparatuses utilizing the phenomenon of attenuation intotal internal reflection such as a surface plasmon resonance sensor ora leaky mode sensor which have been put into practice, there has been aproblem that a long time is required to measure lots of samples. Forexample, when each sample is to be subjected to measurement a pluralityof times at predetermined time intervals, measurement of a second samplecannot be started until measurement of a first sample is finished, whichresults in a very long time required to measure all the samples.

[0023] In view of the foregoing observations and description, a firstobject of the present invention is to provide a measuring apparatusutilizing the phenomenon of attenuation in total internal reflectionwhich can measure lots of samples in a short time.

[0024] A second object of the present invention is to provide ameasuring method utilizing the phenomenon of attenuation in totalinternal reflection which makes it feasible to measure lots of samplesin a short time in the case where each sample is to be subjected tomeasurement a plurality of times at time intervals.

[0025] Further, in conventional surface plasmon resonance sensors, therehas been a problem that measurements can greatly fluctuate when lightbeam in the form of convergent light is caused to enter the dielectricblock in the aforesaid way (1) in order to obtain various angles ofincidence of the light beam to the interface of the dielectric block andthe metal film. For example, the fluctuation in the measurements isdetected as fluctuation in the position of the dark line describedabove.

[0026] The similar problem is recognized also in conventional leaky modesensors when light beam is caused to enter the dielectric block in theaforesaid way (1) in order to obtain various angles of incidence of thelight beam to the interface of the dielectric block and the clad layer.

[0027] Thus a third object of the present invention is to preventgeneration of great fluctuation in the measurements and improve themeasuring accuracy in the apparatuses utilizing the phenomenon ofattenuation in total internal reflection where the light beam is causedto enter the dielectric block in the form of convergent light.

[0028] A fourth object of the present invention is to improve themeasuring accuracy in a measuring method utilizing the phenomenon ofattenuation in total internal reflection.

SUMMARY OF THE INVENTION

[0029] A first measuring apparatus utilizing the phenomenon ofattenuation in total internal reflection for accomplishing the firstobject of the present invention comprises

[0030] a plurality of measuring units each comprising a dielectricblock, a film layer which is formed on a surface of the dielectric blockand a sample holder which holds a sample on the surface of the filmlayer,

[0031] a support which supports the measuring units,

[0032] a light source which emits a light beam,

[0033] an optical system which causes the light beam to enter thedielectric block at various angles of incidence so that the totalinternal reflection condition is satisfied at the interface of thedielectric block and the film layer,

[0034] a photodetector which detects attenuation in total internalreflection by detecting the intensity of the light beam which isreflected in total internal reflection at the interface, and

[0035] a drive means which moves the support relatively to the opticalsystem and the photodetector to bring the measuring units in sequence toa predetermined position with respect to the optical system and thephotodetector where the total internal reflection condition is satisfiedat the interface of the dielectric block and the film layer and variousangles of incidence to the interface can be obtained.

[0036] A second measuring apparatus utilizing the phenomenon ofattenuation in total internal reflection for accomplishing the firstobject of the present invention comprises

[0037] a plurality of measuring units each comprising a dielectricblock, a film layer which is formed on a surface of the dielectricblock, sensing material which interacts a specific component in a sampleand is disposed on the surface of the film layer, and a sample holderwhich holds the sample on the surface of the sensing material,

[0038] a support which supports the measuring units,

[0039] a light source which emits a light beam,

[0040] an optical system which causes the light beam to enter thedielectric block at various angles of incidence so that the totalinternal reflection condition is satisfied at the interface of thedielectric block and the film layer,

[0041] a photodetector which detects attenuation in total internalreflection by detecting the intensity of the light beam which isreflected in total internal reflection at the interface, and

[0042] a drive means which moves the support relatively to the opticalsystem and the photodetector to bring the measuring units in sequence toa predetermined position with respect to the optical system and thephotodetector where the total internal reflection condition is satisfiedat the interface of the dielectric block and the film layer and variousangles of incidence to the interface can be obtained.

[0043] A third measuring apparatus utilizing the phenomenon ofattenuation in total internal reflection for accomplishing the firstobject of the present invention is arranged to measure utilizingespecially the phenomenon of attenuation in total internal reflection bythe aforesaid surface plasmon resonance and comprises

[0044] a plurality of measuring units each comprising a dielectricblock, a metal film layer which is formed on a surface of the dielectricblock and a sample holder which holds a sample on the surface of themetal film layer,

[0045] a support which supports the measuring units,

[0046] a light source which emits a light beam,

[0047] an optical system which causes the light beam to enter thedielectric block at various angles of incidence so that the totalinternal reflection condition is satisfied at the interface of thedielectric block and the metal film layer,

[0048] a photodetector which detects attenuation in total internalreflection by surface plasmon resonance by detecting the intensity ofthe light beam which is reflected in total internal reflection at theinterface, and

[0049] a drive means which moves the support relatively to the opticalsystem and the photodetector to bring the measuring units in sequence toa predetermined position with respect to the optical system and thephotodetector where the total internal reflection condition is satisfiedat the interface of the dielectric block and the metal film layer andvarious angles of incidence to the interface can be obtained.

[0050] A fourth measuring apparatus utilizing the phenomenon ofattenuation in total internal reflection for accomplishing the firstobject of the present invention is also arranged to measure utilizingespecially the phenomenon of attenuation in total internal reflection bythe aforesaid surface plasmon resonance and comprises

[0051] a plurality of measuring units each comprising a dielectricblock, a metal film layer which is formed on a surface of the dielectricblock, sensing material which interacts a specific component in a sampleand is disposed on the surface of the metal film layer, and a sampleholder which holds the sample on the surface of the sensing material,

[0052] a support which supports the measuring units,

[0053] a light source which emits a light beam,

[0054] an optical system which causes the light beam to enter thedielectric block at various angles of incidence so that the totalinternal reflection condition is satisfied at the interface of thedielectric block and the metal film layer,

[0055] a photodetector which detects attenuation in total internalreflection by surface plasmon resonance by detecting the intensity ofthe light beam which is reflected in total internal reflection at theinterface, and

[0056] a drive means which moves the support relatively to the opticalsystem and the photodetector to bring the measuring units in sequence toa predetermined position with respect to the optical system and thephotodetector where the total internal reflection condition is satisfiedat the interface of the dielectric block and the metal film layer andvarious angles of incidence to the interface can be obtained.

[0057] A fifth measuring apparatus utilizing the phenomenon ofattenuation in total internal reflection for accomplishing the firstobject of the present invention is arranged to measure utilizingespecially the phenomenon of attenuation in total internal reflection bythe aforesaid excitation of the waveguide mode at an optical waveguidelayer and comprises

[0058] a plurality of measuring units each comprising a dielectricblock, a film layer consisting of a clad layer formed on a surface ofthe dielectric block and an optical waveguide layer formed on the cladlayer, and a sample holder which holds a sample on the surface of thefilm layer,

[0059] a support which supports the measuring units,

[0060] a light source which emits a light beam,

[0061] an optical system which causes the light beam to enter thedielectric block at various angles of incidence so that the totalinternal reflection condition is satisfied at the interface of thedielectric block and the clad layer,

[0062] a photodetector which detects attenuation in total internalreflection by excitation of waveguide mode at the optical waveguidelayer by detecting the intensity of the light beam which is reflected intotal internal reflection at the interface, and

[0063] a drive means which moves the support relatively to the opticalsystem and the photodetector to bring the measuring units in sequence toa predetermined position with respect to the optical system and thephotodetector where the total internal reflection condition is satisfiedat the interface of the dielectric block and the clad layer and variousangles of incidence to the interface can be obtained.

[0064] A sixth measuring apparatus utilizing the phenomenon ofattenuation in total internal reflection for accomplishing the firstobject of the present invention is arranged to measure utilizingespecially the phenomenon of attenuation in total internal reflection bythe aforesaid excitation of the waveguide mode at an optical waveguidelayer and comprises

[0065] a plurality of measuring units each comprising a dielectricblock, a film layer consisting of a clad layer formed on a surface ofthe dielectric block and an optical waveguide layer formed on the cladlayer, sensing material which interacts a specific component in a sampleand is disposed on the surface of the film layer, and a sample holderwhich holds the sample on the surface of the sensing material,

[0066] a support which supports the measuring units,

[0067] a light source which emits a light beam,

[0068] an optical system which causes the light beam to enter thedielectric block at various angles of incidence so that the totalinternal reflection condition is satisfied at the interface of thedielectric block and the clad layer,

[0069] a photodetector which detects attenuation in total internalreflection by excitation of waveguide mode at the optical waveguidelayer by detecting the intensity of the light beam which is reflected intotal internal reflection at the interface, and

[0070] a drive means which moves the support relatively to the opticalsystem and the photodetector to bring the measuring units in sequence toa predetermined position with respect to the optical system and thephotodetector where the total internal reflection condition is satisfiedat the interface of the dielectric block and the clad layer and variousangles of incidence to the interface can be obtained.

[0071] In the measuring apparatuses utilizing the phenomenon ofattenuation in total internal reflection for accomplishing the firstobject of the present invention, for instance, the drive means moves thesupport with the optical system and the photodetector kept stationary.

[0072] In this case, it is preferred that the support be a turntablewhich supports the measuring units about its axis of rotation and thedrive means be arranged to intermittently rotate the turntable. Thesupport may support the measuring units in a row and the drive means maybe arranged to intermittently move the support in the direction of therow of the measuring units.

[0073] Further, the drive means may move the optical system and thephotodetector with the support kept stationary.

[0074] In this case, it is preferred that the support supports themeasuring units in a circle and the drive means intermittently moves theoptical system and the photodetector along the measuring units in acircle. The support may support the measuring units in a row and thedrive means may be arranged to intermittently move the optical systemand the photodetector along the measuring units in a row.

[0075] When the drive means is provided with a roller bearing whichsupports its rotating shaft, it is preferred that the drive means bearranged to rotate the rotating shaft in one direction when a set ofmeasuring units on the support are measured and to rotate the rotatingshaft in the other direction to return the rotating shaft to theoriginal position after the measurement on the measuring units to waitin the original position for measurement on another set of measuringunits.

[0076] Further, in the measuring apparatuses utilizing the phenomenon ofattenuation in total internal reflection for accomplishing the firstobject of the present invention, it is preferred that the measuringunits be connected in a row to form a measuring unit train, and thesupport be arranged to support the measuring unit train.

[0077] Further, in the measuring apparatuses utilizing the phenomenon ofattenuation in total internal reflection for accomplishing the firstobject of the present invention, it is preferred that an automaticsample feeding means which automatically feeds a sample to the sampleholder of each of the measuring units be provided.

[0078] Further, in the measuring apparatuses utilizing the phenomenon ofattenuation in total internal reflection for accomplishing the firstobject of the present invention, it is preferred that the dielectricblock of each measuring unit be fixed to the support while the filmlayer and the sample holder be integrated with each other to form ameasuring chip, and the measuring chip be exchangeable with respect tothe dielectric block.

[0079] In this case, it is preferred that the measuring apparatus beprovided with a measuring chip cassette in which plurality of measuringchips are contained, and a chip supply means which takes out themeasuring chips from the measuring chip cassette one by one and mountseach measuring chip on the dielectric block.

[0080] In another embodiment, the dielectric block, the film layer andthe sample holder of the measuring unit are integrated with each otherto form a measuring chip which is exchangeable with respect to thesupport.

[0081] In this case, it is preferred that the measuring apparatus beprovided with a measuring chip cassette in which plurality of measuringchips are contained, and a chip supply means which takes out themeasuring chips from the measuring chip cassette one by one and mountseach measuring chip on the support.

[0082] It is preferred that the optical system be arranged to cause thelight beam to enter the dielectric block as a convergent light beam or adivergent light beam, and the photodetector be arranged to detect aposition of a dark line which is generated in the light beam reflectedin total internal reflection at the interface due to attenuation intotal reflection.

[0083] Further, it is preferred that the optical system be arranged tocause the light beam to impinge upon the interface in a defocused state.In this case, it is preferred that the beam diameter of the light beamas measured on the interface in the direction of movement of the supportbe at least ten times the mechanical positioning accuracy of thesupport.

[0084] Further, in the measuring apparatuses utilizing the phenomenon ofattenuation in total internal reflection for accomplishing the firstobject of the present invention, it is preferred that

[0085] the measuring units be supported on the upper side of thesupport,

[0086] the light source be arranged to emit the light beam downward fromabove the support,

[0087] the optical system be provided with a reflecting member whichreflects upward the light beam toward the interface.

[0088] Further, in the measuring apparatuses utilizing the phenomenon ofattenuation in total internal reflection for accomplishing the firstobject of the present invention, it is preferred that

[0089] the measuring units be supported on the upper side of thesupport,

[0090] the optical system be arranged to cause the light beam to impingeupon the interface from below the interface,

[0091] the photodetector is positioned above the support with its lightreceiving face directed downward, and

[0092] a reflecting member which reflects the light beam, reflected intotal internal reflection at the interface, upward toward thephotodetector be provided.

[0093] Further, in the measuring apparatuses utilizing the phenomenon ofattenuation in total internal reflection for accomplishing the firstobject of the present invention, it is preferred that

[0094] a temperature control means which maintains the temperature ofthe measuring units at a predetermined temperature before and/or afterthe measuring units are supported by the support be provided.

[0095] Further, in the measuring apparatuses utilizing the phenomenon ofattenuation in total internal reflection for accomplishing the firstobject of the present invention, it is preferred that a stirrer meanswhich stirs the sample held by the sample holder of the measuring unitsupported by the support before attenuation in total internal reflectionis detected be provided.

[0096] Further, in the measuring apparatuses utilizing the phenomenon ofattenuation in total internal reflection for accomplishing the firstobject of the present invention, it is preferred that

[0097] a reference liquid supply means which supplies at least one ofthe measuring units supported by the support with reference liquid whichhas optical properties related to the optical properties of the samplebe provided, and

[0098] data on the state of attenuation in total internal reflection forthe sample obtained by the photodetector be corrected on the basis ofdata on the state of total internal reflection for the reference liquid.

[0099] In the case where the sample comprises a sample materialdissolved in solvent, it is preferred that the reference liquid supplymeans supplies the solvent as the reference liquid.

[0100] Further, it is preferred that the measuring apparatuses utilizingthe phenomenon of attenuation in total internal reflection foraccomplishing the first object of the present invention be provided with

[0101] an identification mark provided on each of the measuring units,

[0102] a reading means which reads out the identification mark from eachof the measuring units subjected to the measurement,

[0103] a sample information input means which inputs information on thesamples fed to the respective measuring units,

[0104] a display means which displays the result of the measurement, and

[0105] a control means which is connected to the display means, thesample information input means and the reading means to store theidentification mark for each measuring unit and the information on thesample fed to the measuring unit correlated to each other and causes thedisplay means to display the result of the measurement on the sampleheld by a measuring unit in correlation with the identification mark forthe measuring unit and the information on the sample fed to themeasuring unit stored correlated to each other.

[0106] In the method for accomplishing the second object of the presentinvention, a measuring apparatus utilizing the phenomenon of attenuationin total internal reflection for accomplishing the first object of thepresent invention is employed, and the method for accomplishing thesecond object of the present invention comprises the steps of

[0107] detecting attenuation in total internal reflection for the sampleheld in first one of the measuring units,

[0108] moving the support relatively to the optical system and thephotodetector to bring a second one of the measuring units to thepredetermined position with respect to the optical system and thephotodetector and detecting attenuation in total internal reflection forthe sample held in the second one of the measuring units, and

[0109] moving the support relatively to the optical system and thephotodetector to bring the first one of the measuring units again to thepredetermined position with respect to the optical system and thephotodetector and detecting again attenuation in total internalreflection for the sample held in the first one of the measuring units.

[0110] A first measuring apparatus utilizing the phenomenon ofattenuation in total internal reflection for accomplishing the thirdobject of the present invention comprises

[0111] a dielectric block,

[0112] a film layer which is formed on one surface of the dielectricblock and is brought into contact with a sample,

[0113] a light source which emits a light beam,

[0114] an optical system which causes the light beam to enter thedielectric block in convergent light so that the total internalreflection condition is satisfied at the interface of the dielectricblock and the film layer and various angles of incidence of the lightbeam to the interface of the dielectric block and the film layer can beobtained, and

[0115] a photodetector which detects attenuation in total internalreflection by detecting the intensity of the light beam which isreflected in total internal reflection at the interface, and

[0116] is characterized in that the optical system is arranged so thatthe light beam is not focused on the interface.

[0117] A second measuring apparatus utilizing the phenomenon ofattenuation in total internal reflection for accomplishing the thirdobject of the present invention is arranged to measure utilizingespecially the phenomenon of attenuation in total internal reflection bythe aforesaid surface plasmon resonance and comprises

[0118] a dielectric block,

[0119] a metal film layer which is formed on one surface of thedielectric block and is brought into contact with a sample,

[0120] a light source which emits a light beam,

[0121] an optical system which causes the light beam to enter thedielectric block in convergent light so that the total internalreflection condition is satisfied at the interface of the dielectricblock and the metal film layer and various angles of incidence of thelight beam to the interface of the dielectric block and the metal filmlayer can be obtained, and

[0122] a photodetector which detects attenuation in total internalreflection by surface plasmon resonance by detecting the intensity ofthe light beam which is reflected in total internal reflection at theinterface, and

[0123] is characterized in that the optical system is arranged so thatthe light beam is not focused on the interface.

[0124] A third measuring apparatus utilizing the phenomenon ofattenuation in total internal reflection for accomplishing the thirdobject of the present invention is in the form of a leaky mode sensorand comprises

[0125] a dielectric block,

[0126] a film layer which is formed of a clad layer formed on a surfaceof the dielectric block and an optical waveguide layer formed on theclad layer and is brought into contact with a sample,

[0127] a light source which emits a light beam,

[0128] an optical system which causes the light beam to enter thedielectric block in convergent light so that the total internalreflection condition is satisfied at the interface of the dielectricblock and the clad layer and various angles of incidence of the lightbeam to the interface of the dielectric block and the clad layer can beobtained, and

[0129] a photodetector which detects attenuation in total internalreflection by excitation of waveguide mode at the optical waveguidelayer by detecting the intensity of the light beam which is reflected intotal internal reflection at the interface, and

[0130] is characterized in that the optical system is arranged so thatthe light beam is not focused on the interface.

[0131] In the measuring apparatuses utilizing the phenomenon ofattenuation in total internal reflection for accomplishing the thirdobject of the present invention, it is preferred that the light beam beat least 500 μm in a cross-sectional size at least in one direction onthe interface (the interface of the dielectric block and the metal filmlayer in the case of the surface plasmon resonance sensor, the interfaceof the dielectric block and the clad layer in the case of the leaky modesensor).

[0132] As the optical system, one which focuses the light beam so thatthe interface is positioned outside the focal depth of the light beamcan be suitably employed. In this case, the focal depth means a range inwhich the beam diameter is within twice that at the focal point.Further, as the optical system, one which is arranged so that the lightbeam is not focused on the interface due to its aberration may also beemployed.

[0133] For example, the optical system may comprise an optical system inwhich the light beam is converged in a conical shape by a spherical lensor an optical system in which the light beam is converged in awedge-like shape by a cylindrical lens.

[0134] In the measuring apparatuses utilizing the phenomenon ofattenuation in total internal reflection for accomplishing the thirdobject of the present invention, it is preferred that a sensing mediumwhich makes bonding reaction with a specific material in the sample befixed on the film layer (the metal film layer in the case of the surfaceplasmon resonance sensor, the optical waveguide layer in the case of theleaky mode sensor). In this specification, the expression “the filmlayer is in contact with the sample” should be broadly interpreted toinclude a state where the film layer is in contact with the sample layerwith such a sensing medium intervening therebetween.

[0135] Further, in the measuring apparatuses utilizing the phenomenon ofattenuation in total internal reflection for accomplishing the thirdobject of the present invention, it is preferred that a sample holderfor holding the sample on the film layer and/or a sample introductionmechanism for introducing the sample onto the film layer be provided.

[0136] A first measuring method utilizing the phenomenon of attenuationin total internal reflection for accomplishing the fourth object of thepresent invention comprises the steps of

[0137] bringing a sample into contact with a film layer formed on onesurface of a dielectric block,

[0138] causing a light beam to enter the dielectric block in convergentlight so that the total internal reflection condition is satisfied atthe interface of the dielectric block and the film layer and variousangles of incidence of the light beam to the interface of the dielectricblock and the film layer can be obtained, and

[0139] detecting attenuation in total internal reflection by detectingthe intensity of the light beam which is reflected in total internalreflection at the interface, and

[0140] is characterized by the step of causing the light beam to enterthe dielectric block not to be focused on the interface.

[0141] A second measuring method utilizing the phenomenon of attenuationin total internal reflection for accomplishing the fourth object of thepresent invention comprises the steps of

[0142] bringing a sample into contact with a metal film layer formed onone surface of a dielectric block,

[0143] causing a light beam to enter the dielectric block in convergentlight so that the total internal reflection condition is satisfied atthe interface of the dielectric block and the metal film layer andvarious angles of incidence of the light beam to the interface of thedielectric block and the metal film layer can be obtained, and

[0144] detecting attenuation in total internal reflection by surfaceplasmon resonance by detecting the intensity of the light beam which isreflected in total internal reflection at the interface, and

[0145] is characterized by the step of causing the light beam to enterthe dielectric block not to be focused on the interface.

[0146] A third measuring method utilizing the phenomenon of attenuationin total internal reflection for accomplishing the fourth object of thepresent invention comprises the steps of

[0147] bringing a sample into contact with an optical waveguide layerformed on a clad layer formed on one surface of a dielectric block,

[0148] causing a light beam to enter the dielectric block in convergentlight so that the total internal reflection condition is satisfied atthe interface of the dielectric block and the clad layer and variousangles of incidence of the light beam to the interface of the dielectricblock and the clad layer can be obtained, and

[0149] detecting attenuation in total internal reflection by detectingthe intensity of the light beam which is reflected in total internalreflection at the interface, and

[0150] is characterized by the step of causing the light beam to enterthe dielectric block not to be focused on the interface.

[0151] It is preferred that the light beam is caused to enter thedielectric block so as to be at least 500 μm in a cross-sectional sizeat least in one direction on the interface.

DISCLOSURE OF THE INVENTION

[0152] In the measuring apparatuses utilizing the phenomenon ofattenuation in total internal reflection for accomplishing the firstobject of the present invention, since a plurality of measuring unitseach comprising a dielectric block, a film layer (a metal film layer inthe case of that utilizing surface plasmon resonance, a clad layer andan optical waveguide layer in the case of that utilizing excitation ofwaveguide mode), and the support is moved relatively to the opticalsystem and the photodetector to bring the measuring units in sequence toa predetermined position with respect to the optical system and thephotodetector, a plurality of measuring units carrying thereon samplescan be subjected to the measurement in sequence, whereby lots of samplescan be measured in a short time.

[0153] For example, when the support is a turntable which supports themeasuring units about its axis of rotation and the drive means isarranged to intermittently rotate the turntable, or the support supportsthe measuring units in a row and the drive means is arranged tointermittently move the support in the direction of the row of themeasuring units, measurement on lots of samples can be carried out athigh efficiency.

[0154] Further, also when the support supports the measuring units in acircle and the drive means is arranged to intermittently rotate theoptical system and the photodetector along the measuring units, or thesupport supports the measuring units in a row and the drive means isarranged to intermittently move the optical system and the photodetectoralong the row of the measuring units, measurement on lots of samples canbe carried out at high efficiency.

[0155] In the measuring apparatuses utilizing the phenomenon ofattenuation in total internal reflection for accomplishing the firstobject of the present invention in which sensing material whichinteracts a specific component in a sample is held on the surface of thefilm layer, the state of attenuation in total internal reflection, thatis, the state of surface plasmon resonance or the state of excitation ofwaveguide mode, is changed by the interaction, and accordingly, thespecific reaction between the specific material in the sample and thesensing material can be detected by detecting the change of the state ofattenuation in total internal reflection.

[0156] When the drive means is provided with a roller bearing whichsupports its rotating shaft, and the drive means is arranged to rotatethe rotating shaft in one direction when a set of measuring units on thesupport are measured and to rotate the rotating shaft in the otherdirection to return the rotating shaft to the original position afterthe measurement on the measuring units to wait in the original positionfor measurement on another set of measuring units, the angular positionof the rollers of the roller bearing when a measuring unit on a givenposition of the support is brought to the predetermined position isconstant, whereby deterioration in measuring accuracy due to fluctuationin the angular position of the rollers can be prevented.

[0157] Further, when the measuring units are connected in a row to forma measuring unit train and the support is arranged to support themeasuring unit train, the measuring units can be accurately located inplace and becomes easier to handle, which results in higher measuringefficiency.

[0158] Further, when the sample holder of each of the measuring units isprovided with an automatic sample feeding means which automaticallyfeeds a sample to the measuring unit, the time required to feed thesample to the measuring unit can be shortened, and lots of samples canbe measured in a further shorter time.

[0159] Further, when the dielectric block of each measuring unit isfixed to the support while the film layer and the sample holder isintegrated with each other to form a measuring chip, and the measuringchip is exchangeable with respect to the dielectric block, new samplescan be subjected to measurement in sequence by replacing the measuringunits which have been measured with measuring units loaded with othersamples, whereby lots of samples can be measured in a further shortertime.

[0160] In this case, when a measuring chip cassette in which pluralityof measuring chips are contained and a chip supply means which takes outthe measuring chips from the measuring chip cassette one by one andmounts each measuring chip on the dielectric block are employed, supplyof the measuring chips can be effectively carried out, whereby lots ofsamples can be measured in a further shorter time.

[0161] When the dielectric block, the film layer and the sample holderof the measuring unit are integrated with each other to form a measuringchip which is exchangeable with respect to the support, new samples canalso be subjected to measurement in sequence by replacing the measuringunits which have been measured with measuring units loaded with othersamples, whereby lots of samples can be measured in a further shortertime.

[0162] Also in this case, when a measuring chip cassette in whichplurality of measuring chips are contained and a chip supply means whichtakes out the measuring chips from the measuring chip cassette one byone and mounts each measuring chip on the dielectric block are employed,supply of the measuring chips can be effectively carried out, wherebylots of samples can be measured in a further shorter time.

[0163] In the case where the support is mechanically moved by the drivemeans, fluctuation in position of the support is inevitable. Howeverfluctuation in position of the support results in fluctuation inposition of the measuring unit with respect to the optical system, whichresults in an error in measuring attenuation in total internalreflection. For example, an error in detecting the position of the darkline representing attenuation total internal reflection can occur. As acause for leading fluctuation in position of the support to an error inmeasuring attenuation in total internal reflection, fluctuation in thethickness of the film layer, the thickness of the sensing material layerand/or the reacting weight of the sensing material and the samplematerial by position is conceivable.

[0164] When the optical system is arranged to cause the light beam toimpinge upon the interface in a defocused state, errors in detecting thestate of attenuation in total internal reflection (e.g., detection ofthe position of the dark line) are averaged and as a result, themeasuring accuracy is improved.

[0165] The measuring accuracy can be further higher when the beamdiameter of the light beam as measured on the interface in the directionof movement of the support be at least ten times the mechanicalpositioning accuracy of the support. The reason is as follows. That is,in this case, since the positioning error is {fraction (1/10)} of thebeam diameter at most and the remainder {fraction (9/10)} is constantlyincluded in the measuring range, the signal error generated due to thepositioning error can be suppressed to {fraction (1/10)}, which ispractically negligible in the normal quantitative analysis.

[0166] In the measuring apparatuses utilizing the phenomenon ofattenuation in total internal reflection for accomplishing the firstobject of the present invention, when the measuring units are supportedon the upper side of the support, the light source is arranged to emitthe light beams downward from above the support, the optical system isprovided with a reflecting member which reflects upward the light beamtoward the interface, it becomes unnecessary to take into accountinterference between the support and the optical system including thelight source, and the freedom in layout of the optical system and thelight source as well as other elements which are to be disposed near thesupport can be increased.

[0167] Further, when the measuring units are supported on the upper sideof the support, the optical system is arranged to cause the light beamto impinge upon the interface from below the interface, thephotodetector is positioned above the support with its light receivingface directed downward, and a reflecting member which reflects the lightbeam, reflected in total internal reflection at the interface, upwardtoward the photodetector is provided, it becomes unnecessary to takeinto account interference between the support and the photodetector, andthe freedom in layout of the photodetector as well as other elementswhich are to be disposed near the support can be increased.

[0168] When the measuring apparatuses utilizing the phenomenon ofattenuation in total internal reflection for accomplishing the firstobject of the present invention is provided with a temperature controlmeans which maintains the temperature of the measuring units at apredetermined temperature before and/or after the measuring units aresupported by the support, deterioration in measuring accuracy due tochange of temperature of the sample in the measuring unit can beprevented.

[0169] When the measuring apparatuses utilizing the phenomenon ofattenuation in total internal reflection for accomplishing the firstobject of the present invention is provided with a stirrer means whichstirs the sample held by the sample holder of the measuring unitsupported by the support before attenuation in total internal reflectionis detected, deterioration in measuring accuracy due to unevenconcentration distribution of the sample material in the sample liquidcan be prevented.

[0170] Further, in the measuring apparatuses utilizing the phenomenon ofattenuation in total internal reflection for accomplishing the firstobject of the present invention, when a reference liquid supply meanswhich supplies at least one of the measuring units supported by thesupport with reference liquid which has optical properties related tothe optical properties of the sample is provided, and data on the stateof attenuation in total internal reflection for the sample obtained bythe photodetector is corrected on the basis of data on the state oftotal internal reflection for the reference liquid, change of therefractive index of the solvent of the sample, for instance, with changeof the environmental temperature or the like, change of the propertiesof the optical system, for instance, with change of the environmentaltemperature, and the like can be compensated for and the properties ofthe sample material in the sample liquid can be correctly measured.

[0171] Further when the measuring apparatuses utilizing the phenomenonof attenuation in total internal reflection for accomplishing the firstobject of the present invention is provided with

[0172] an identification mark provided on each of the measuring units,

[0173] a reading means which reads out the identification mark from eachof the measuring units subjected to the measurement,

[0174] a sample information input means which inputs information on thesamples fed to the respective measuring units,

[0175] a display means which displays the result of the measurement, and

[0176] a control means which is connected to the display means, thesample information input means and the reading means to store theidentification mark for each measuring unit and the information on thesample fed to the measuring unit correlated to each other and causes thedisplay means to display the result of the measurement on the sampleheld by a measuring unit in correlation with the identification mark forthe measuring unit and the information on the sample fed to themeasuring unit stored correlated to each other,

[0177] since the identification mark for each measuring unit, theinformation on the sample fed to the measuring unit and the result ofthe measurement are managed in correlation with each other, an eventthat measurement is done in a wrong combination of a measuring unit andthe sample liquid or the result of measurement for a wrong sample isdisplayed can be prevented.

[0178] In the method for accomplishing the second object of the presentinvention, since after attenuation in total internal reflection for thesample held in first one of the measuring units is detected, the supportis moved relatively to the optical system and the photodetector to bringa second one of the measuring units to the predetermined position withrespect to the optical system and the photodetector, attenuation intotal internal reflection for the sample held in the second one of themeasuring units is detected, and then the support is moved relatively tothe optical system and the photodetector to bring the first one of themeasuring units again to the predetermined position with respect to theoptical system and the photodetector to detect again attenuation intotal internal reflection for the sample held in the first one of themeasuring units, measurement on one sample material can be carried outbetween intervals of measurement on another sample material, wherebylots of samples can be efficiently measured in a shorter time.

[0179] Investigation by these inventors has revealed that the problem,inherent to the prior art, that the measured values greatly fluctuate isdue to the fact that the light beam is focused on the interface of thedielectric block and the film layer.

[0180] That is, when the light beam is focused on the interface, thespot size of the light beam on the interface becomes as very small as 10μm to several hundreds of μm. Whereas there are fine irregularities onthe surface of the film layer such as a metal film layer, and at thesame time, when the specific material in the sample liquid is caused toreact with the sensing material fixed to the surface of the film layer,reaction properties fluctuate by the position of the sensing material.Accordingly, when the light beam impinges upon the interface in a finespot, the measured value is greatly affected by the fine irregularitieson the surface of the film layer and the reaction properties which arelargely vary according to the position of the light beam, which resultsin large fluctuation in the measured values.

[0181] On the basis of this recognition, in the measuring apparatusesutilizing the phenomenon of attenuation in total internal reflection foraccomplishing the third object of the present invention, the opticalsystem is arranged so that the light beam is not focused on theinterface. With this arrangement, the spot size of the light beam on theinterface becomes larger than in the conventional apparatuses. When thespot size of the light beam on the interface is large, the measuredvalues come to correspond to the average irregularity on the surface ofthe film layer and the average reaction property, whereby fluctuation inthe measured values can be suppressed.

BRIEF DESCRIPTION OF THE DRAWINGS

[0182]FIG. 1 is a perspective view showing a surface plasmon resonancesensor in accordance with a first embodiment of the present invention,

[0183]FIG. 2 is a side view partly cut away showing an important part ofthe surface plasmon resonance sensor shown in FIG. 1,

[0184]FIG. 3 is a graph showing the relation between the angle ofincidence of the light beam and the output of the photodetector in thesurface plasmon resonance sensor,

[0185]FIG. 4 is a side view partly cut away showing an important part ofa surface plasmon resonance sensor in accordance with a secondembodiment of the present invention,

[0186]FIG. 5 is a side view partly cut away showing an important part ofa surface plasmon resonance sensor in accordance with a third embodimentof the present invention,

[0187]FIG. 6 is a side view showing a surface plasmon resonance sensorin accordance with a fourth embodiment of the present invention,

[0188]FIG. 7 is a plan view showing the surface plasmon resonance sensorin accordance with the fourth embodiment of the present invention,

[0189]FIG. 8 is a side view partly cut away showing an important part ofthe surface plasmon resonance sensor in accordance with the fourthembodiment of the present invention,

[0190]FIG. 9 is a side view partly cut away showing an important part ofa leaky mode sensor in accordance with a fifth embodiment of the presentinvention,

[0191]FIG. 10 is a side view partly cut away showing an important partof a leaky mode sensor in accordance with a sixth embodiment of thepresent invention,

[0192]FIG. 11 is a side view showing a surface plasmon resonance sensorin accordance with a seventh embodiment of the present invention,

[0193]FIG. 12 is a side view showing a surface plasmon resonance sensorin accordance with an eighth embodiment of the present invention,

[0194]FIG. 13 is a side view partly cut away showing an example of thestructure of the drive shaft portion of a turntable which can beemployed in the surface plasmon resonance sensor of the presentinvention,

[0195]FIG. 14 is a side view partly cut away showing an important partof a surface plasmon resonance sensor in accordance with a ninthembodiment of the present invention,

[0196]FIG. 15 is a block diagram showing an electrical arrangement ofthe surface plasmon resonance sensor shown in FIG. 14,

[0197]FIGS. 16A to 16C are views showing the relation between the angleof incidence of the light beam and the detected intensity of light, andthe relation between the angle of incidence of a light beam and thedifferential value of the light intensity signal in the surface plasmonresonance sensor shown in FIG. 14,

[0198]FIGS. 17A and 17B are graphs for illustrating an example of changein the measured values in the surface plasmon resonance sensor shown inFIG. 14,

[0199]FIG. 18 is a plan view showing a surface plasmon resonance sensorin accordance with a tenth embodiment of the present invention,

[0200]FIG. 19 is a side view partly cut away showing an important partof the surface plasmon resonance sensor shown in FIG. 18,

[0201]FIG. 20 is a side view partly cut away showing an important partof a surface plasmon resonance sensor in accordance with an eleventhembodiment of the present invention,

[0202]FIG. 21 is a perspective view partly cut away showing an importantpart of a surface plasmon resonance sensor in accordance with a twelfthembodiment of the present invention,

[0203]FIG. 22 is a perspective view showing a surface plasmon resonancesensor in accordance with a thirteenth embodiment of the presentinvention,

[0204]FIG. 23 is a perspective view showing a part of the surfaceplasmon resonance sensor shown in FIG. 22,

[0205]FIG. 24 is a plan view showing a part of the surface plasmonresonance sensor shown in FIG. 22,

[0206]FIG. 25 is a front view showing a part of the surface plasmonresonance sensor shown in FIG. 22,

[0207]FIG. 26 is a perspective view showing an example of a measuringunit train employed in the surface plasmon resonance sensor of thepresent invention,

[0208]FIG. 27 is a perspective view showing another example of ameasuring unit train employed in the surface plasmon resonance sensor ofthe present invention,

[0209]FIG. 28 is a fragmentary side view of a surface plasmon resonancesensor in accordance with a fourteenth embodiment of the presentinvention,

[0210]FIG. 29 is a schematic view showing a surface plasmon resonancesensor in accordance with a fifteenth embodiment of the presentinvention,

[0211]FIG. 30 is a view for illustrating the flow of control andinformation processing in the surface plasmon resonance sensor shown inFIG. 29,

[0212]FIG. 31 is a perspective view showing an important part of asurface plasmon resonance sensor in accordance with a sixteenthembodiment of the present invention,

[0213]FIG. 32 is a side view partly cut away of another example of themeasuring unit employed in the surface plasmon resonance sensor of thepresent invention,

[0214]FIG. 33 is a side view partly cut away of a leaky mode sensor inaccordance with a seventeenth embodiment of the present invention,

[0215]FIGS. 34A to 34G are views for illustrating the flow of control ina surface plasmon resonance measuring method in accordance with anembodiment of the present invention, and

[0216]FIG. 35 is a graph showing an example of result of measurement bya surface plasmon resonance measuring method of the present invention.

PREFERRED EMBODIMENTS OF THE INVENTION

[0217] Embodiments of the present invention will be described in detailwith reference to the drawings, hereinbelow. FIG. 1 shows the overallarrangement of a surface plasmon resonance sensor in accordance with afirst embodiment of the present invention, and FIG. 2 is a side viewshowing an important part of the surface plasmon resonance sensor.

[0218] As shown in FIG. 1, the surface plasmon resonance sensorcomprises a turn table 20 on which a plurality of measuring units 10 aresupported, a laser 31 such as a semiconductor laser which emits ameasuring light beam (laser beam) 30, a condenser lens 32 which forms anincident optical system, a photodetector 40, a drive means 50 whichintermittently rotates the turn table 20, a controller 60 which controlsthe drive means 50 and at the same time receives output signal S of thephotodetector 40 to execute processing described later, and an automaticsample supply mechanism 70.

[0219] As shown in FIG. 2, each of the measuring units 10 comprises atransparent dielectric block 11 which is, for instance, rectangular inshape, a metal film 12 such as of gold, silver, cupper, aluminum or thelike which is formed on the upper face of the block 11, and a sampleholder frame 13 which is a tubular member defining above the metal film12 a space having closed side walls. In the sample holder frame 13 isstored, for instance, a sample liquid 15 in the manner to be describedlater.

[0220] The measuring unit 10 is formed, for instance, by integrallyforming the dielectric block 11 and the sample holder frame 13 byinjection molding of transparent resin and is replaceable. In thisparticular embodiment, the measuring units 10 are removably held inthrough holes formed in the turn table 20. Further, in this particularembodiment, a sensing medium 14 is fixed on the metal film 12. Thesensing medium 14 will be described later.

[0221] The turn table 20 is arranged so that a plurality of (eleven inthis particular embodiment) measuring units 10 are supported on a circleabout its axis of rotation 20 a at regular intervals. The drive means 50comprises a stepping motor or the like and intermittently rotates theturn table 20 by an angle equal to the angular space between the units10.

[0222] As shown in FIG. 2, the condenser lens 32 condenses the lightbeam 30 and caused to enter the dielectric block 11 in the form of aconvergent light beam so that various angles of incidence of the lightbeam 30 to the interface 11 a of the dielectric block 11 and the metalfilm 12 can be obtained. The range of the angles of incidence is set toinclude a range in which the total internal reflection condition of thelight beam 30 is satisfied at the interface 11 a and a surface plasmonresonance can take place.

[0223] The light beam 30 impinges upon the interface 11 a in the form ofp-polarized light. This can be realized by suitably positioning thelaser 31 or by controlling the polarization direction of the light beam30 with a wavelength plate, a polarizing plate or the like.

[0224] The photodetector 40 comprises a line sensor having a number ofphotodetector elements arranged in a row in the direction arrow X inFIG. 2.

[0225] The controller 60 receives an address signal A representing theposition of the drive means 50 and outputs a drive signal D for drivingthe drive means 50 on the basis of a predetermined sequence. Further,the controller 60 is provided with a signal processing section 61 whichreceives an output signal S from the photodetector 40 and a displaysection 62 which receives an output of the signal processing section 61.

[0226] The automatic sample supply mechanism 70 comprises a pipette 71which sucks a predetermined amount of sample liquid and a pipette drivemeans 72 which moves the pipette 71. The automatic sample supplymechanism 70 causes the pipette 72 to suck the sample in a samplecontainer 73 held in a predetermined position and moves the pipette 71above the sample holder frame 13 of a measuring unit 10 in apredetermined position, and causes the pipette 71 to spot the sampleliquid in the sample holder frame 13.

[0227] Operation of the surface plasmon resonance sensor will bedescribed hereinbelow. The turn table 20 is intermittently rotated bythe drive means 50. When the turn table 20 is stopped, sample liquid 15is supplied to the sample holder frame 13 of the measuring unit 10 inthe predetermined position by the automatic sample supply mechanism 70.

[0228] When a measuring unit 10 the sample holder frame 13 of whichcontains therein the sample liquid 15 is stopped in a predeterminedmeasuring position (the position of the right side measuring unit 10 inFIG. 2), the laser 31 is driven under the control of the controller 60and the light beam 30 emitted from the laser 31 impinges upon theinterface 11 a between the dielectric block 11 and the metal film 12 inthe form of convergent light. The light beam 30 is reflected in totalinternal reflection at the interface 11 a and is detected by thephotodetector 40.

[0229] Since the light beam 30 impinges upon the interface 11 a in theform of convergent light, the light beam 30 includes components whichimpinge upon the interface at various angles θ. The angles θ ofincidence is not smaller than the angle of total internal reflection.Accordingly, the light beam 30 is reflected in total internal reflectionat the interface 11 a, and the reflected light beam 30 at the interface11 a includes components reflected at various reflecting angles.

[0230] When the light beam 30 is reflected in total internal reflectionat the interface 11 a, evanescent waves ooze out from the interface 11 atoward the metal film 12. The component of the light beam 30 whichimpinges upon the interface at a specified angle θsp resonates with thesurface plasmon excited on the surface of the metal film 12 by theevanescent waves and accordingly, the intensity I of the component ofthe light beam 30 reflected in total internal reflection at theinterface 11 a sharply drops. FIG. 3 is a graph showing the relationbetween the angle of incidence θ of the light beam 30 and the intensityI of the reflected light.

[0231] By detecting the amount of reflected light received by eachphotodetector element on the basis of the output signal S of thephotodetector 40, the position of the photodetector element whichdetects the dark line can be determined and the angle θsp of incidenceat which the intensity I of the light beam 30 reflected in totalinternal reflection at the interface 11 a (angle θsp of total internalreflection attenuation) sharply drops can be determined. Accordingly byreferring to a standard curve which represents the relation between theintensity I of the reflected light and the angle θ of incidence of thelight beam 30, a particular material in the sample 15 can bequantitatively analyzed. The signal processing section 61 of thecontroller 60 quantitatively analyzes the particular material in thesample 15 on the basis of this fact and the result of the analysis isdisplayed by the display section 62.

[0232] When each sample 15 is subjected to measurement only once, themeasuring units 10 may be manually or automatically removed from theturn table 20. Whereas when each sample 15 is subjected to measurement aplurality of times, the measuring units 10 may be left on the turn table20 so that the measuring unit 20 is brought to the measuring positionagain when the turn table 20 makes another rotation.

[0233] As described above, in the surface plasmon resonance sensor ofthis embodiment, since a plurality of measuring units 10 are held on theturn table 20 and the turn table 20 is intermittently rotated so thatthe measuring units 10 are brought to the measuring position insequence, a plurality of samples 15 can be measured at a high efficiencywhereby the time required to measure each sample can be shortened.

[0234] Further, since, in this particular embodiment, the automaticsample supply mechanism 70 is provided, the time required to supplysamples to the measuring units 10 also can be shortened, whereby thetime required to measure each sample can be further shortened.

[0235] Further, since, in this particular embodiment, the dielectricblock 11, the metal film layer 12 and the sample holder frame 13 areintegrated into a measuring unit 10, new samples 15 can be measured insequence by changing measuring units 10 after the measurement with ameasuring unit 10, whereby the time required to measure a lot of samplescan be further shortened.

[0236] The sensing medium 14 fixed on the surface of the metal film 12is selectively linked to a specific material in the sample 15. As such acombination of the sensing medium 14 and the specific material in thesample 15, for instance, combinations of antigen and antibody are known.In such a case, antigen-antibody reaction can be detected on the basisof the angle θsp of total internal reflection attenuation.

[0237] That is, the refractive index of the sensing medium 14 changeswith progress of bonding reaction between the specific material in thesample and the sensing medium 14 so that the curve shown in FIG. 3 movesright and left. Accordingly, the state of antigen-antibody reaction canbe detected through the total reflection attenuation angle θsp. In thiscase, both the sample 15 and the sensing medium 14 can be analyzed.

[0238] A surface plasmon resonance sensor in accordance with a secondembodiment of the present invention will be described with reference toFIG. 4, hereinbelow. FIG. 4 is a side view partly cut away showing animportant part of a surface plasmon resonance sensor in accordance witha second embodiment of the present invention. In FIG. 4, elementsanalogous to those shown in FIG. 2 are given the same reference numeralsand will not be described here. (and so forth)

[0239] The surface plasmon resonance sensor shown in FIG. 4 differs fromthat shown in FIG. 2 in the structure of the measuring unit. That is,the measuring unit 10′ in this embodiment comprises a dielectric block11′ fixed to the turntable 20, and a sample holder frame 13′ and a metalfilm layer 12 integrated with each other. The sample holder frame 13′ isin the form of a bottomed tubular member formed of transparentdielectric material and the metal film layer 12 is fixed on the bottomof the sample holder frame 13′. Thus, the sample holder frame 13′ andthe metal film layer 12 are integrated to form a measuring chip.

[0240] The measuring chip can be removed from the dielectric block 11′so that the measuring chip can be replaced by another. It is preferredthat refractive index matching fluid is provided between the sampleholder frame 13′ and the dielectric block 11′. In this case, the sampleholder frame 13′ is integrated with the dielectric block 11′ to form asingle dielectric block and the light beam 30 is caused to impinge uponthe interface 13 a between the sample holder frame 13′ and the metalfilm layer 12.

[0241] Also in this embodiment, by removing the measuring chip, bearingthereon the sample which has been measured, from the dielectric blockand replacing with another, new samples 15 can be measured in sequence,whereby the time required to measure a lot of samples can be furthershortened.

[0242] A surface plasmon resonance sensor in accordance with a thirdembodiment of the present invention will be described with reference toFIG. 5, hereinbelow. Figure is a side view partly cut away showing animportant part of the surface plasmon resonance sensor in accordancewith the third embodiment of the present invention. The surface plasmonresonance sensor shown in FIG. 5 differs from that shown in FIG. 2 inthe manner of projecting the light beam 30 onto the interface 11 a ofthe dielectric block 11 and the metal film layer 12. That is, in thisembodiment, the condenser lens 32 is arranged so that the light beam 30(in the form of a conical beam) impinges upon the interface 11 a in adefocused state. The light beam 30 is 500 μm×500 μm in spot size on theinterface 11 a, whereas, in the conventional system where the light beamis focused on the interface 11 a, the light beam 30 is 100 μm×100 μm inspot size on the interface 11 a.

[0243] Since the turntable 20 is mechanically driven by the drive means50, fluctuation in position of the turntable 20 is inevitable.Fluctuation in position of the turntable 20 results in fluctuation inposition of the measuring unit 10 with respect to the light beam 30emanating from the condenser lens 32, which results in an error inmeasuring attenuation in total internal reflection. For example, anerror in detecting the position of the dark line representingattenuation total internal reflection can occur.

[0244] When the light beam 30 impinges upon the interface 11 a in adefocused state, errors in detecting the state of surface plasmonresonance (in detecting the position of the aforesaid dark line) areaveraged, whereby the measuring accuracy is enhanced.

[0245] When the spot size is large as in this embodiment, the measuredvalue reflects the average of the fine irregularities on the surface ofthe metal film layer 12 and the reaction properties, whereby fluctuationof the measured values can be suppressed.

[0246] When the light beam 30 is caused to impinge upon the interface 11a in a defocused state, it is preferred that the beam diameter of thelight beam 30 as measured on the interface 11 a in the direction ofmovement of the turntable 30 be at least ten times the mechanicalpositioning accuracy of the turntable 30.

[0247] A fourth embodiment of the present invention will be describedwith reference to FIGS. 6, 7 and 8. FIGS. 6 and 7 respectively show aside view and a plan view of a surface plasmon resonance sensor inaccordance with the fourth embodiment of the present invention, and FIG.8 is a side view showing an important of the same.

[0248] As shown in FIGS. 6 and 7, in the surface plasmon resonancesensor of this embodiment, four measuring units 80 are supported on aturntable 20 at regular intervals of 90°, and the turntable 20 isintermittently rotated 90° by 90° in the direction of arrow R. Thus,each measuring unit 80 is brought to four positions, a measuring unitsupply position P1, a sample supply position P2, a measuring position P3and a measuring unit discharge position P4, in sequence. The measuringunit 80 will be described in detail later.

[0249] When a measuring unit support portion is stopped in the measuringunit supply position P1, a measuring chip supply means 76 takes out ameasuring unit 80 from a cassette 75 in which a plurality of measuringunits 80 are contained and supplies it to the measuring unit supplyportion. The chip supply means 76 may be of a known structure comprisinga suction cup and a mechanism for moving the suction cup. For example,the suction cup is arranged to take out the measuring units 80 one byone through a take-out port formed in the bottom of the cassette 75holding the measuring unit 80 under suction force and the mechanismmoves the suction cup together with the measuring unit 80 to themeasuring unit support portion in the measuring unit supply position P1.

[0250] To the measuring unit stopped in the sample supply position P2, asample is supplied by an automatic sample supply mechanism 70.

[0251] The sample held by the measuring unit 80 stopped in the measuringposition P3 is analyzed by a surface plasmon resonance sensing means 77.The analysis will be described in detail later with reference to FIG. 8.

[0252] The measuring unit 80 stopped in the measuring unit dischargeposition P4 is discharged from the turntable 20 by a measuring chipdischarge means 78. The measuring unit support position thus removedwith the measuring unit is brought to the measuring unit supply positionP1 as the turntable 20 is subsequently turned by 90° and supplied withanother measuring unit 80. Thereafter, the aforesaid steps are repeatedeach time the turntable 20 is turned by 90°.

[0253] Analysis by the surface plasmon resonance sensing means 77 willbe described with reference to FIG. 8, hereinbelow. The measuring unit80 employed in this embodiment comprises a transparent electric block81, a metal film layer 82 and a sample holder frame 83 which are formedintegrally with each other as in the measuring unit 10 employed in thefirst embodiment.

[0254] A laser beam 30 emitted from the laser 31 is condensed by acondenser lens 90 and is reflected by a mirror 91 to impinge upon theinterface 81 a of the dielectric block 81 and the metal film layer 82.The light beam 30 reflected in total internal reflection at theinterface 81 a is reflected by a mirror 92 to impinge upon thephotodetector 40 after collimated by a collimator lens 93. The outputsignal S of the photodetector 40 is input into the signal processingsection 61 of the controller 60 (FIG. 6) and the sample material isanalyzed on the basis of the output signal S in the same manner as thatdescribed above in conjunction with the first embodiment.

[0255] As can be understood from the description above, in thisembodiment, since a plurality of measuring chips (measuring units) 80are contained in the measuring chip cassette 75, and the measuring chips80 are taken out from the measuring chip cassette 75 one by one andmounted on the turntable 20 by the chip supply means 76, supply of themeasuring chips can be effectively carried out, whereby lots of samples15 can be measured in a shorter time.

[0256] This arrangement can be applied to not only the measuring units80 each of which comprises a transparent dielectric block 81, a metalfilm layer 82 and a sample holder frame 83 integrally formed with eachother but also measuring units 10′ in which a metal film layer 12 and asample holder frame 13′ are formed to be exchangeable with respect tothe dielectric block 11′ as shown in FIG. 4 by integrating the metalfilm layer 12 and the sample holder frame 13′ into a measuring chip andcontaining a plurality of measuring chips in a cassette, whereby themeasuring chips can be automatically supplied and supply of themeasuring chips can be effectively carried out.

[0257] Though, in the embodiments described above, a turntable 20 isemployed as the support for supporting the measuring units, the supportneed not be limited to such a turntable. For example, the support may beone which is linearly moved back and forth with a plurality of measuringunits supported thereon to bring the measuring units to the measuringposition one by one.

[0258] In this case, when each sample is subjected to measurement aplurality of times, a plurality of measuring means each comprising anoptical system for projecting a light beam onto the measuring unit and aphotodetector are provided along the support so that each measuring unitis brought to one of the measuring means as the support is moved.Otherwise, each sample can be subjected to measurement a plurality oftimes by providing a single measuring means along the support andreversing the support after a certain number of measuring units aresubjected to measurement by moving the support in the regular direction.

[0259] Also the support in the form of the aforesaid turntable 20 may berotated back and forth to subject each measuring unit to measurement.Further it is possible to provide a plurality of measuring means alongthe turntable 20 so that each measuring unit is subjected to measurementa plurality of times during one rotation of the turntable 20.

[0260] A fifth embodiment of the present invention will be described,hereinbelow. FIG. 9 is a side view showing a measuring apparatusutilizing the phenomenon of attenuation in total internal reflection inaccordance with the fifth embodiment of the present invention. In FIG.9, elements analogous to those shown in FIG. 2 are given the samereference numerals.

[0261] The measuring apparatus of this embodiment is a leaky mode sensorand measuring units 110 in the form of a measuring chip are employed inthis embodiment. Each measuring unit 110 comprises a dielectric block11, a clad layer 111 formed on one surface (the upper surface as seen inFIG. 9) and an optical waveguide layer 112 formed on the clad layer 111.

[0262] The dielectric block 11 is formed of, for instance, syntheticresin or optical glass such as BK7. The clad layer 111 is formed ofdielectric material lower than the dielectric block 1 in refractiveindex or metal such as gold. The optical wave guide layer 112 is formedof dielectric material such as PMMA which is higher than the clad layer111 in refractive index. The thickness of the clad layer 111 is 36.5 nmwhen it is formed of gold film, and the thickness of the opticalwaveguide layer 112 is about 700 nm when it is formed of PMMA.

[0263] In the leaky mode sensor, when the light beam 30 emitted from thelaser 31 is caused to impinge upon the clad layer 111 at an angle notsmaller than an angle of total internal reflection through thedielectric block 11, the light beam 30 is reflected in total internalreflection at the interface 11 a of the dielectric block 11 and the cladlayer 111, and only light having a particular wave number and impingingupon the waveguide layer 112 at a particular angle of incidence comes topropagate through the optical waveguide layer 112 in a waveguide modeafter passing through the clad layer 111. When the waveguide mode isthus excited, almost all the incident light is taken in the opticalwaveguide layer 112 and accordingly, the intensity of light reflected intotal internal reflection at the interface 11 a of the dielectric block11 and the clad layer 111 sharply drops. That is, attenuation in totalinternal reflection occurs.

[0264] Since the wave number of light to be propagated through theoptical waveguide layer 112 in a waveguide mode depends upon therefractive index of the sample 15 on the optical waveguide layer 112,the refractive index and/or the properties of the sample 15 related tothe refractive index can be detected on the basis of the angle ofincidence at which the attenuation in total internal reflection occurs.A signal processing section 61 quantitatively analyzes the particularmaterial in the sample 15 on the basis of this fact and the result ofthe analysis is displayed by the display section (not shown).

[0265] Also in the leaky mode sensor of this embodiment, since aplurality of measuring units 110 are held on the turn table 20 and theturn table 20 is intermittently rotated so that the measuring units 110are brought to the measuring position in sequence, a plurality ofsamples 15 can be measured at a high efficiency whereby the timerequired to measure each sample can be shortened.

[0266] Further, since, also in this particular embodiment, thedielectric block 11, the clad layer 111 and the optical waveguide layer112 are integrated into a measuring unit 110, new samples 15 can bemeasured in sequence by changing measuring units 110 after themeasurement with a measuring unit 110, whereby the time required tomeasure a lot of samples can be further shortened.

[0267] A sixth embodiment of the present invention will be described,hereinbelow. FIG. 10 is a side view showing an important part of ameasuring apparatus utilizing the phenomenon of attenuation in totalinternal reflection in accordance with the sixth embodiment of thepresent invention. In FIG. 10, elements analogous to those shown in FIG.9 are given the same reference numerals.

[0268] The measuring apparatus of this embodiment is also a leaky modesensor and measuring units 120 in the form of a measuring chip areemployed in this embodiment. Each measuring unit 120 comprises adielectric block 11, a clad layer 111 formed on one surface (the uppersurface as seen in FIG. 9) of the dielectric block 11, an opticalwaveguide layer 112 formed on the clad layer 111 and a sensing medium 14fixed on the optical waveguide layer 112. The apparatus of thisembodiment basically differs from that shown in FIG. 9 only in that thesensing material 14 is fixed on the optical waveguide layer 112.

[0269] The sensing medium 14 fixed on the surface of the metal film 12is selectively linked to a specific material in the sample 15 as that ofthe apparatus shown in FIG. 2. As such a combination of the sensingmedium 14 and the specific material in the sample 15, for instance,combinations of antigen and antibody are known. In such a case,antigen-antibody reaction can be detected on the basis of the angle θspof total internal reflection attenuation.

[0270] That is, though the relation between the angle of incidence θ ofthe light beam 30 and the intensity I of the reflected light isbasically as shown in FIG. 3, the effective refractive index of thesensing medium 14 changes with progress of bonding reaction between thespecific material in the sample and the sensing medium 14 so that thecurve shown in FIG. 3 moves right and left. Accordingly, the state ofantigen-antibody reaction can be detected through the total reflectionattenuation angle θsp.

[0271] A seventh embodiment of the present invention will be described,hereinbelow. FIG. 11 is a side view showing an important part of ameasuring apparatus utilizing the phenomenon of attenuation in totalinternal reflection in accordance with the seventh embodiment of thepresent invention. In FIG. 11, elements analogous to those shown inFIGS. 1 and 2 are given the same reference numerals.

[0272] The measuring apparatus of this embodiment is a surface plasmonresonance sensor and is arranged so that a measuring unit 10 containingtherein a sample 15 is held on a measuring unit holding portion 128. Themeasuring unit 10 is substantially the same as that shown in FIGS. 1 and2 except that the shape of the dielectric block 11 somewhat differs fromthat of the dielectric block shown in FIGS. 1 and 2.

[0273] In the apparatus of this embodiment, though the light beam 30 iscaused to impinge upon the interface 11 a of the dielectric block 11 andthe metal film layer 12 from below as in the embodiment shown in FIG. 2,a mirror 121 which reflects upward the light beam 30 reflected in totalinternal reflection at the interface 11 a is provided and the light beam30 reflected by the mirror 121 to travel upward is detected by aphotodetector 40 which is disposed above the mirror 121 with its lightreceiving surface directed downward.

[0274] When the photodetector 40 is positioned above the turntable 20 byproviding a mirror 121 which reflects upward the light beam 30, itbecomes unnecessary to take into account interference between therotating turntable 20 and the photodetector 40, and the freedom inlayout of the photodetector 40 as well as other elements which are to bedisposed near the support can be increased.

[0275] An eighth embodiment of the present invention will be described,hereinbelow. FIG. 12 is a side view showing an important part of ameasuring apparatus utilizing the phenomenon of attenuation in totalinternal reflection in accordance with the eighth embodiment of thepresent invention. The measuring apparatus of this embodiment is also asurface plasmon resonance sensor and differs from that shown in FIG. 11in that a mirror 122 which reflects upward the light beam 30 emittedfrom the laser 31 and the laser 31 is disposed above the mirror 122.

[0276] By thus disposing the photodetector 40 and the laser 31 above theturntable 20, it becomes unnecessary to take into account interferencebetween the turntable 20 and the photodetector 40 as well as the laser31, and the freedom in layout of the photodetector 40 and the laser 31as well as other elements which are to be disposed near the turntable 20can be increased.

[0277] The layout of the photodetector 40 and the laser 31 employed inthe seventh and eighth embodiments of the present invention need not belimited to the surface plasmon resonance sensor but can be employed alsoin the leaky mode sensor shown in FIG. 9 or 10.

[0278] Further, the layout of the photodetector 40 and the laser 31employed in the seventh and eighth embodiments of the present inventionneed not be limited to the surface plasmon resonance sensor or the leakymode sensor where a turntable is employed as the support but can beemployed also in the surface plasmon resonance sensor or the leaky modesensor where a support is linearly moved back and forth.

[0279]FIG. 13 shows in more detail the turntable 20 shown in FIG. 1.That is, the turntable 20 is operatively connected to a motor 50 a of asupport drive means 50 such as a stepping motor by way of a rotatingshaft 20 a and is driven by the motor 50 a. The rotating shaft 20 a issupported for rotation on a bearing portion 50 b of the support drivemeans 50 by way of one or more roller bearing 130.

[0280] The roller bearing 130 comprises an inner ring 130 a which isfitted on the rotating shaft 20 a and fixed thereto, an outer ring 130 bwhich is fitted in the bearing portion 50 b of the support drive means50 and fixed thereto, and rolling members 130 c such as rollers or ballswhich roll between the inner and outer rings 130 a and 130 b. When therotating shaft 20 is rotated by the motor 50 a, the inner ring 130 a isrotated integrally with the rotating shaft 20 a and the rolling members130 c make revolution around the inner ring 130 a while rotating on theinner ring 130 a.

[0281] Accordingly, when the rolling members 130 are not good inroundness and/or the surface of the inner ring 130 a or the outer ring130 b is rough, the position in a diametrical direction or theinclination of the inner ring 130 a (that is, of the rotating shaft 20a) varies according to the revolution position of the rolling members130 c, which can result in fluctuation in the axis of rotation of theturntable 20 and/or inclination of the turntable 20 from the horizon.Such a behavior of the turntable 20 changes the position of themeasuring units 10 thereon and the incident angle of the light beam 30to the interface 11 a fluctuates, whereby error in measuring the samples15 is generated.

[0282] In order to prevent production of the measuring error, rotationof the turntable 20 is controlled in the following manner. Here it isassumed that sixteen measuring units 10 are supported on the turntable20 at regular intervals of 22.5°. The sixteen positions on the turntable20 in which the sixteen measuring units 10 are held will be respectivelyreferred to as 1ch (channel) to 16ch.

[0283] The measuring unit 10 held in each channel is subjected tomeasurement by the measuring system shown in FIG. 2 each time it isstopped in the measuring position.

[0284] In this particular embodiment, the measuring units 10 in channelsat intervals of 112.5° or at intervals of five channels are brought tothe measuring position in sequence. That is, the measuring units 10 in1ch, 6ch, 11ch and 16ch are brought to the measuring position insequence in this order during a first rotation of the turntable in theregular direction, the measuring units 10 in 5ch, 10ch and 15ch arebrought to the measuring position in sequence in this order during asecond rotation of the turntable in the same direction, the measuringunits 10 in 4ch, 9ch and 14ch are brought to the measuring position insequence in this order during a third rotation of the turntable in thesame direction, the measuring units 10 in 3ch, 8ch and 13ch are broughtto the measuring position in sequence in this order during a fourthrotation of the turntable in the same direction, and the measuring units10 in 2ch, 7ch and 12ch are brought to the measuring position insequence in this order during a fifth rotation of the turntable in thesame direction.

[0285] While one of the measuring units 10 is stopped in the measuringposition and is subjected to measurement, other measuring units 10stopped in the respective positions are subjected to operationscorresponding to the positions, e.g., supply of a sample 15.

[0286] After the turntable 20 is intermittently rotated by fiverotations and the measuring units 10 in all the channels are subjectedto predetermined operations, the turntable 20 is continuously rotated byfive rotations in the reverse direction. While the turntable 20 isrotated by five rotations in the regular direction, the rolling members130 c make revolution around the inner ring 130 a and when the turntable20 is rotated by five rotations in the reverse direction, each rollingmember 130 c is returned to the original revolution position.

[0287] Accordingly, when change with time of the properties shown inFIG. 3 is to be detected through a plurality of measurements on onesample 15, each measurement on one sample 15 can be done with therolling members 130 c held in the same revolution position by thusreturning the rolling members 130 c in the original position after eachsample 15 is subjected to measurement during five rotations of theturntable 20 in the regular direction. With this arrangement, eachmeasuring unit 10 can be held in the same position for all themeasurements, whereby production of measuring errors due to fluctuationin the incident angle of the light beam 30 to the interface 11 a frommeasurement to measurement.

[0288] The number of channels on the turntable 20, the angle by whichthe turntable 20 is intermittently rotated, the number of rotations ofthe turntable 20 for one measurement of the respective measuring units10, and the like may be set freely without limited to those describedabove.

[0289] The control of the turntable 20 described above may be alsoapplied to the leaky mode sensor shown in FIG. 9 or 10 without limitedto the surface plasmon resonance sensor.

[0290] The measured values by the surface plasmon resonance sensor orthe leaky mode sensor is apt to change according to environmentalconditions such as the environmental temperature. This is because, forinstance, the refractive index of the sample liquid changes with thetemperature and the properties of the measuring optical system finelychange with the temperature. A ninth embodiment of the present inventionwhere change of the measured value due to change in the environmentalconditions is prevented will be described, hereinbelow.

[0291]FIG. 14 is a side view showing an important part of a measuringapparatus utilizing the phenomenon of attenuation in total internalreflection in accordance with the ninth embodiment of the presentinvention. The measuring apparatus of this embodiment is also a surfaceplasmon resonance sensor and comprises a dielectric block 210 which issubstantially a quadrangular pyramid in shape, and a metal film layer212 of gold, silver, copper, aluminum or the like formed on one surface(the upper surface as seen in FIG. 14) of the dielectric block 210.

[0292] The dielectric block 210 is formed of, for instance, transparentresin and is thickened at a part 210 a circumscribing the metal filmlayer 212. The thickened part 210 a of the dielectric block 210functions as a sample holder containing therein a sample liquid 211. Inthis embodiment, a sensing medium 230 (which will be described later) isfixed on the metal film layer 212.

[0293] The dielectric block 210 forms together with the metal film layer212 a disposable measuring unit (measuring chip) 222 and the disposablemeasuring units 222 are held in chip holding holes 231 a formed in aturntable 231 which is the same as the turntable 20 shown in FIG. 1.With the dielectric blocks 210 are thus supported on the turntable 231,the turntable 231 is intermittently rotated predetermined angle bypredetermined angle, and a sample liquid 211 is spotted onto thedielectric block 210 stopped in a predetermined position to be held inthe sample holder 210 a. When the turntable 231 is subsequently rotatedby a predetermined angle, a dielectric block 210 is stopped in themeasuring position shown in FIG. 14.

[0294] The surface plasmon resonance sensor of this embodiment isprovided with, in addition to the dielectric block 210, a laser 214 suchas a semiconductor laser which emits a light beam 213, an incidentoptical system 215 which causes the light beam 213 to enter thedielectric block 210 so that various angles of incidence of the lightbeam 213 to the interface 210 b of the dielectric block 210 and themetal film layer 212 can be obtained, a collimator lens 16 whichcollimates the light beam 213 reflected in total internal reflection atthe interference 210 b into a parallel light beam, a photodetector 17which detects the parallel light beam, a differential amplifier array 18connected to the photodetector 17, a driver 19, a signal processingsection 220 which may comprise, for instance, a computer system, and adisplay means 21 connected to the signal processing section 220.

[0295]FIG. 15 shows the electrical arrangement of the surface plasmonresonance sensor. As shown in FIG. 15, the driver 19 comprises sampleholding circuits 22 a, 22 b, 22 c . . . which respectively sample-holdthe outputs of the differential amplifiers 18 a, 18 b, 18 c . . . of thedifferential amplifier array 18, a multiplexer 23 into which the outputsof the sample-holding circuits 22 a, 22 b, 22 c . . . , an A/D convertor24 which digitizes the output of the multiplexer 23 and inputs digitalsignals into a signal processing section 220, a drive circuit 25 whichdrives the multiplexer 23 and the sample-holding circuits 22 a, 22 b, 22c . . . , and a controller 26 which controls the drive circuit 25 underthe control of the signal processing section 220.

[0296] As shown in FIG. 14, the light beam 213 emanating from the laser214 in the form of a divergent light beam is converged on the interface210 b of the dielectric block 210 and the metal film layer 212 by theoptical system 215. Accordingly, the light beam 213 includes componentsimpinging upon the interface 210 b at various angles of incidence θ,which is not smaller than the total internal reflection angle.Accordingly, the light beam 213 is reflected in total internalreflection at the interface 210 b, and the light beam 213 reflected atthe interface 210 b includes components reflected at various angles.

[0297] The light beam 213 is caused to impinge upon the interface 210 bin the form of p-polarized light. This can be realized by positioningthe laser 214 so that the light beam 213 is polarized in the suitabledirection or by controlling the polarization direction of the light beam213 with a wavelength plate, a polarizing plate or the like.

[0298] After reflected in total internal reflection at the interface 210b and collimated into a parallel light beam by the collimator lens 16,the light beam 213 is detected by the photodetector 17. In thisparticular embodiment, the photodetector 17 is a photodiode arraycomprising a plurality of photodiodes 17 a, 17 b, 17 c . . . arranged ina row, and is disposed so that the photodiode array extends inperpendicular to the direction of travel of the parallel light beam 213in the plane of FIG. 14. Accordingly, components of the light beam 213reflected at different angles at the interface 210 b are received bydifferent photodiodes 17 a, 17 b, 17 c . . . .

[0299] The outputs of the photodiodes 17 a, 17 b, 17 c . . . are inputinto the differential amplifiers 18 a, 18 b, 18 c . . . of thedifferential amplifier array 18, and at this time, the outputs ofadjacent two photodiodes are input into one differential amplifiers.Accordingly, the outputs of the differential amplifiers 18 a, 18 b, 18 c. . . may be considered to be values obtained by differentiatingphotodetecting signals output from the photodiodes 17 a, 17 b, 17 c . .. in the direction which the photodiode array extends.

[0300] The outputs of the differential amplifiers 18 a, 18 b, 18 c . . .. are sample-held by the sample-holding circuits 22 a, 22 b, 22 c . . .. at predetermined timings and input into the multiplexer 23. Themultiplexer 23 inputs the sampled outputs of the differential amplifiers18 a, 18 b, 18 c . . . into the A/D convertor 24 in a predeterminedorder. The A/D convertor 24 digitizes the sampled outputs and inputs thedigitized signals into the signal processing section 220.

[0301]FIGS. 16A to 16C are views illustrating the relation between theintensity I of the reflected light beam 213 reflected at the interface210 b versus the angle of incidence q FIGS. 16A to 16C are viewsillustrating the relation between the intensity I of the reflected lightbeam 213 reflected at the interface 210 b versus the angle of incidenceθ of the light beam 213 to the interface 210 b and the outputs of thedifferential amplifiers 18 a, 18 b, 18 c . . . . It is assumed that therelation between the intensity I of the reflected light beam 213reflected at the interface 210 b versus the angle of incidence θ of thelight beam 213 to the interface 210 b is as shown in FIG. 16A.

[0302] Light impinging upon the interface 210 b at a particular angle ofincidence θ_(sp) generates surface plasmon on the interface between themetal film layer 212 and the sample 211 and accordingly, for lightimpinging upon the interface 210 b at a particular angle of incidenceθ_(sp), the intensity I of light reflected in total internal reflectionat the interface 210 b sharply drops. That is, the angle θ_(sp) is thetotal reflection attenuation angle and the intensity I takes a minimumvalue at the angle θ_(sp). The drop of the intensity I is observed as adark line in the reflected light as denoted by D in FIG. 14.

[0303]FIG. 16B shows the direction in which the photodiodes 17 a, 17 b,17 c . . . are arranged and the positions of the photodiodes 17 a, 17 b,17 c . . . in the direction are in one to one correspondence with theangles of incidence θ.

[0304] The relation between the positions of the photodiodes 17 a, 17 b,17 c . . . in the direction in which the photodiodes 17 a, 17 b, 17 c .. . are arranged, that is, the angles of incidence θ, and the outputs I′of the differential amplifiers 18 a, 18 b, 18 c . . . (differentialvalues of the intensity I of the reflected light) is as shown in FIG.16C.

[0305] The signal processing section 220 selects a differentialamplifier whose output is the closest to or equal to 0 (the differentialamplifier 18 d in the example shown in FIG. 16C) and causes the displaymeans 21 to display the differential value I′ output from the selecteddifferential amplifier after carrying out correction processing to bedescribed later on the differential value I′.

[0306] Thereafter, each time a predetermined time lapses, a differentialvalue I′ output from the selected amplifier 18 d is displayed aftersubjected to the correction processing.

[0307] Since the differential value I′ increases or decreases inresponse to right and left movement of the curve shown in FIG. 16A,which represents change of the dielectric constant or the refractiveindex of the material in contact with the metal film layer 212 (FIG. 14)of the measuring unit 222. Accordingly, by continuously measuring thedifferential value I′, the change of the refractive index or theproperties of the material in contact with the metal film layer 212 canbe detected.

[0308] Especially, in this embodiment, since the aforesaid sensingmedium 230 which combines with a specific material (sample material) inthe sample liquid 211 is fixed on the metal film later 212, and therefractive index of the sensing medium 230 changes with the state ofbonding of the sensing medium 230 and the specific material, change ofthe state of bonding of the sensing medium 230 and the specific materialcan be detected by continuously measuring the differential value I′. Inthis case, both the sample liquid 211 and the sensing medium 230 can beanalyzed. As such a combination of the sensing medium 230 and thespecific material, for instance, combinations of antigen and antibodyare known.

[0309] As can be understood from the description above, since aphotodiode array comprising a plurality of photodiodes 17 a, 17 b, 17 c. . . arranged in a row is employed as the photodetector 17, the darkline can be detected even if the curve shown in FIG. 16A largely movesright and left according to the kind of the sample liquid 211, whereby alarge dynamic range of measurement can be ensured.

[0310] Instead of a differential amplifier array 18 comprising aplurality of differential amplifiers 18 a, 18 b, 18 c . . . , a singledifferential amplifier may be employed. In this case, outputs of thephotodiodes 17 a, 17 b, 17 c . . . are switched by a multiplexer so thatoutputs of pairs of adjacent photodiodes are input into the singledifferential amplifier in sequence.

[0311] The aforesaid correction processing to be carried out on thedifferential value I′ will be described in detail, hereinbelow. As shownin FIG. 14, an automatic reference liquid supply mechanism 221 isprovided above a position in which the measuring unit 222 on theturntable 231 is stopped. The structure of the automatic referenceliquid supply mechanism 221 is basically the same as that of the sampleliquid supply mechanism 70 shown in FIG. 1 and the automatic referenceliquid supply mechanism 221 supplies one of the measuring units 222 onthe turntable 231 with the reference liquid. The measuring unit 222supplied with the reference liquid is supplied with no sample liquid.

[0312] The sample liquid 211 in this embodiment comprises a samplematerial dissolved in solvent, and as the reference liquid, the solventis employed. For example, when the sample material is Insulin-BiotinLabeled, and the solvent is PBS (phosphoric acid buffer) containingtherein 0.1% BSA solution, PBS containing therein 0.1% BSA solution isemployed as the reference liquid.

[0313] With one of the measuring units 222 on the turntable 231 suppliedwith the reference liquid and the other measuring units 222 suppliedwith respective sample liquids 211, the differential values I′ of allthe measuring units 222 including the measuring unit supplied with thereference liquid are measured.

[0314]FIG. 17A shows change with time of the differential value I′ on asample liquid 211 and that on a reference liquid, and FIG. 17B showschange with time of the differential value I′ on another sample liquid211 and that on another reference liquid. In FIGS. 17A and 17B, changewith time of the differential value I′ on the sample liquid 211 is shownby hutched circles (measured data) and change with time of thedifferential value I′ on the reference liquid is shown by white circles(correction data).

[0315] As described above, the state of bonding of the sensing medium230 and the specific material in the sample liquid 211 can be determinedby detecting change with time of the measured data, that is, thedifferential value I′ on the sample liquid 211. However, the measureddata is affected by environmental conditions such as the environmentaltemperature since, for instance, the refractive index of the sampleliquid 211 changes with the temperature and the like, and the opticaldistances between elements of the measuring optical system finely changewith the temperature and the like.

[0316] The signal processing section 220 corrects the measureddifferential value I′ by subtracting the value of the correction datafrom the measured differential value I′. That is, the corrected data isas shown by the solid line FIGS. 17A and 17B. The subtraction is madebetween the sample liquid and the reference liquid which are the same inreaction time with the sensing medium 230.

[0317] The differential value I′ on the reference liquid depends uponthe refractive index of the reference liquid, i.e., the solvent of thesample liquid 211 irrespective of the specific material (samplematerial) in the sample liquid 211, and also reflects fine displacementof elements of the optical system, if any. Accordingly, by subtractingthe differential value I′ on the reference liquid from the measuringdata, that is, the differential value I′ on the sample liquid 211,change of the refractive index of the solvent and/or change of theproperties of the optical system can be compensated for, whereby thecorrected data can purely represent properties of the sample material inthe sample liquid 211.

[0318] A tenth embodiment of the present invention will be described,hereinbelow. FIG. 18 is a plan view showing an important part of ameasuring apparatus utilizing the phenomenon of attenuation in totalinternal reflection in accordance with the tenth embodiment of thepresent invention. The measuring apparatus of this embodiment is asurface plasmon resonance sensor and in FIG. 18, elements analogous tothose shown in FIG. 7 are given the same reference numerals. That is,reference numeral 20 denotes a turntable which is intermittently rotatedin the direction of arrow R, reference numeral 70 denotes an automaticsample supply mechanism, reference numeral 76 denotes a chip supplymeans, reference numeral 77 denotes a surface plasmon resonance sensingmeans and reference numeral 78 denotes a measuring chip discharge means.

[0319] In this particular embodiment, the turntable 20 supports 96measuring units 80 and is intermittently rotated 75° by 75° (=360/96).The chip supply means 76 transfers the measuring units 80 on awell-plate 240, containing therein 96 measuring units 80, set in awell-plate setting station 241, one by one to the turntable 20. In thisembodiment, a sensing medium 14 similar to that shown in FIG. 2 is fixedon the metal film layer of each of the measuring units 80, and eachmeasuring unit 80 is supplied to the turntable 20 in the state.

[0320] The sample liquids 15 are stored in wells of 96 well plates 242and the sample supply mechanism 70 transfers the samples 15 in thewell-plates 242 set in a well-plate setting station 243 one by one tothe measuring units 80 on the turntable 20. Then the surface plasmonresonance sensing means 77 carried out measurement on the respectivemeasuring units 80 in the manner described above. After the measurement,the measuring units 80 are discharged from the turntable 20 to adischarge well-plate 244 by the measuring chip discharge means 78.

[0321] The apparatus with the arrangement described above can besuitably employed in random screening for finding a specific materialcombined with a predetermined sensing material as described above.

[0322] As described above, measured values by the surface plasmonresonance sensor vary with change of the refractive index of the sampleliquid due to change of the environmental temperature. An arrangementfor suppressing change of the temperature of the sample liquid 15 willbe described, hereinbelow.

[0323]FIG. 19 is a side view of the well-plate setting station 243 ofthe automatic sample supply mechanism 70. As shown in FIG. 19, there isprovided in the well-plate setting station 243 a metal block 245 having96 recesses 245 a which receives the wells (portions for holding thesample liquid 15) of the well plate 242. A temperature control means 246which may comprise, for instance, a Peltier element or a heater forchanging the temperature of the metal block 245 and a temperature sensor247 are connected to the metal block 245. The temperature detectingsignal S1 of the temperature sensor 247 is input into a controller 248and the controller 248 controls the temperature control means 246 on thebasis of the temperature detecting signal S1 to keep the temperature ofthe metal block 245 at a predetermined temperature. It is preferred thatthe metal block 245 be formed of a metal having a high heat conductivitysuch as copper.

[0324] When the metal block 245 is thus kept at the predeterminedtemperature, the sample liquid 15 in the wells of the well-plate 242 canbe kept also at the predetermined temperature, whereby change of therefractive index of the sample liquid 15 can be prevented andfluctuation of the measured values by the surface plasmon resonancesensor can be prevented.

[0325] The metal 245 may be kept at the predetermined temperature bycirculating temperature-controlled liquid instead of using a temperaturecontrol means 246 such as a Peltier element of a heater as in a surfaceplasmon resonance sensor in accordance with an eleventh embodiment ofthe present invention shown in FIG. 20.

[0326] That is, in the surface plasmon resonance sensor shown in FIG.20, the metal block 245 is provided with a flow passage 245 b foruniformly circulating liquid through the metal block 245. A circulatingpassage 250 is connected to the upstream and downstream ends of the flowpassage 245 b. A pump 251 for flowing liquid such as water underpressure and a heater 252 for heating the liquid are provided in thecirculating passage 250.

[0327] The heater 252 is controlled by a temperature sensor 247 and acontroller 248 which are similar to those shown in FIG. 19 so that theliquid flowing through the flow passage 245 b is kept at a predeterminedtemperature and accordingly, the metal block 245 is kept at apredetermined temperature. Thus, change of the refractive index of thesample liquid 15 can be prevented and fluctuation of the measured valuesby the surface plasmon resonance sensor can be prevented.

[0328] In view of preventing fluctuation of the measured values due tochange in the temperature of the sample liquid 15, it is preferred thatchange of the temperature of the sample liquid 15 supplied to themeasuring units 80 on the turntable 20 be further suppressed. A surfaceplasmon resonance sensor in accordance with a twelfth embodiment of thepresent invention, in which change of the temperature of the sampleliquid 15 supplied to the measuring units 80 on the turntable 20 isfurther suppressed, will be described, hereinbelow.

[0329]FIG. 21 is a perspective view showing an important part of asurface plasmon resonance sensor in accordance with the twelfthembodiment of the present invention. In FIG. 21, elements analogous tothose shown in FIGS. 18 and 19 are given the same reference numerals.

[0330] In this embodiment, the upper side of the turntable 20 is coveredwith a cover 260 so that air just above the turntable 20 is confinedinside the cover 260 and air communication between the inside and theoutside of the cover 260 becomes difficult. The cover 260 is a lowsubstantially cylindrical member with a closed upper end. The cover 260is provided with a small opening 260 a of a size just enough to receivea sample liquid spotting pipette 71 of the automatic sample supplymechanism 70, an opening 260 b of a size just enough to receive thesurface plasmon resonance sensing means 77 comprising a casing 262holding therein the optical system of the sensing means 77, a smallopening 260 c for receiving a chip supply means (not shown), and a smallopening 260 d for receiving a chip discharge means (not shown).

[0331] On the cover 260 are mounted a temperature control means 246which may comprise, for instance, a Peltier element or a heater forchanging the temperature inside the cover 260 and a temperature sensor247 which detects the temperature inside the cover 260. The temperaturedetecting signal S1 of the temperature sensor 247 is input into acontroller 248 and the controller 248 controls the temperature controlmeans 246 on the basis of the temperature detecting signal S1 to keepthe temperature inside the cover 260 at a predetermined temperature.

[0332] When the temperature inside the cover 260 is thus kept at thepredetermined temperature, the sample liquid 15 in the measuring units80 on the turntable 20 can be kept also at the predeterminedtemperature, whereby change of the refractive index of the sample liquid15 can be prevented and fluctuation of the measured values by thesurface plasmon resonance sensor can be prevented.

[0333] Both the mechanism for keeping the temperature of the sampleliquid 15 in the measuring units 80 on the turntable 20 at a presettemperature described above and the mechanism for keeping thetemperature of the sample liquid 15 before supplied to the measuringunits 80 at the preset temperature shown in FIG. 19 or 20 may beemployed. With this arrangement, fluctuation of the measured values bythe surface plasmon resonance sensor due to change in temperature can bemore effectively prevented.

[0334] The mechanism for keeping the temperature of the sample liquid 15may be applied also to the leaky mode sensor shown in FIG. 9 or 10 inorder to prevent fluctuation of the measured values due to change intemperature.

[0335] When the sample liquid 15 is of a type where a sample material isdissolved in solvent, it is desired for the sample liquid 15 to be wellstirred to uniform the concentration of the sample material beforemeasurement in order to correctly analyze properties of the samplematerial. This is true also of leaky mode sensors shown in FIGS. 9 and10.

[0336] In order to well stir the sample liquid 15, for instance, in theapparatus shown in FIG. 21, the turntable 20 may be rotated in onedirection at a high speed or may be alternately rotated in one directionand the other before measurement with the measuring units 80 supportedthereon.

[0337] Otherwise, the sample liquid 15 may be stirred by, after eachsample liquid 15 is spotted into the measuring unit 80 by the pipette71, dipping the pipette 71 in the sample liquid 15 and causing thepipette 71 to repeat sucking the sample liquid 15 and discharging thesame a plurality of times.

[0338] Though the embodiments described above employ a turntable as thesupport for supporting measuring units, embodiments of the presentinvention employing a support which is linearly moved back and forthwill be described hereinbelow.

[0339]FIG. 22 is a perspective view showing a surface plasmon resonancesensor in accordance with a thirteenth embodiment of the presentinvention. In FIG. 22, elements analogous to those shown in FIG. 2 aregiven the same reference numerals.

[0340] In the surface plasmon resonance sensor of this embodiment, asthe support for supporting a plurality of measuring units 10 is employeda slide block 401 which is fitted on a pair of parallel guide rods 400to be linearly slidable in the direction of arrow Y. The slide block 401is in mesh with a precision screw 402 parallel to the guide rods 400.The precision screw 402 is rotated in the regular and reverse directionsby a pulse motor 403, which forms together with the precision screw 402a support drive means.

[0341] The pulse motor 403 is driven under the control of a motorcontroller 404. That is, an output signal S40 of a linear encoder (notshown) which is incorporated in the slide block 401 to detect theposition of the slide block 401 in the longitudinal direction of theguide rods 400 is input into the motor controller 404, and the motorcontroller 404 controls the pulse motor 403 on the basis of the outputsignal S40.

[0342] A laser 31 and a condenser lens 32 are provided on one side ofthe guide rods 400 and a photodetector 40 is disposed on the oppositeside of the guide rods 400. The laser 31, the condenser lens 32 and thephotodetector 40 are basically the same as those shown in FIG. 2.

[0343] In this embodiment, comprising stick-like measuring unit train410 comprising eight measuring units 10 fixed together is employed, andeight measuring units in a row are set on the slide block 401. FIG. 23shows in detail the structure of the measuring unit train 410. As shownin FIG. 23, the measuring unit train 410 comprises eight measuring units10, each being the same as that shown in FIG. 2, connected together by aconnecting member 411.

[0344] For example, a plurality of (twelve in this particularembodiment) the measuring unit trains 410 are set on a plate 420, andthe measuring units 10 are transported and handled in this state. Thatis, the measuring units are placed on a plate 420 by 96 pieces andtransported and handled on the plate.

[0345] When a plurality of measuring units 10 are handled in one pieceas a measuring unit train 410, it is preferred that a dispensingmechanism 430 such as shown in FIG. 25 be employed as the means forsupplying the sample liquid to the measuring units 10. The dispensingmechanism 430 comprises a plurality of dispensing nozzles 431 the numberof which is equal to the number of the measuring units 10 of themeasuring unit train 410 and which are supported on a support member 432at the same pitches as those at which the measuring units 10 areconnected in the measuring unit train 410. The dispensing mechanism 430can supply the sample liquid 15 to all the measuring units 10 of onemeasuring unit train 410 at one time, whereby sample supply can beefficiently accomplished.

[0346] When the surface plasmon resonance sensor shown in FIG. 22 isoperated, one of the measuring unit trains 410 on the plate 420 shown inFIG. 24 is set to a unit setting portion 401 a of the slide block 401 bya supply means (not shown). As shown in FIG. 22, the slide block 401 isa frame-like member having a plurality of through holes 401 b extendingthrough the slide block 401 in the horizontal direction (verticaldirection as seen in FIG. 22), the light beam 30 emitted from the laser31 can be projected onto the measuring unit 10 supported on the slideblock 401 through the through hole 401 b and the light beam 30 reflectedin total internal reflection at the interface of the dielectric block 11and the metal film layer 12 can be detected by the photodetector 40 thethrough hole 401 b.

[0347] When surface plasmon resonance is measured by the apparatus ofthis embodiment, the pulse motor 403 is driven to move the slide block401 to bring a first measuring unit 10, that is, the rightmost measuringunit 10 in FIG. 22, to a measuring position where the measuring unit isexposed to the light beam 30. Then surface plasmon resonance of thefirst measuring unit 10 is measured in the manner described above.

[0348] After the measurement on the first measuring unit 10, the pulsemotor 403 is driven to move the slide block 401 by a distance equal tothe pitch of the measuring units 10 to bring a second measuring unit 10to the measuring position. Surface plasmon resonance of the secondmeasuring unit 10 is then measured. In this manner, intermittentmovement of the slide block 401 and measurement of surface plasmonresonance are repeated until all the eight measuring units 10 aresubjected to measurement.

[0349] Thereafter, the pulse motor 403 is rotated in the reversedirection to return the slide block 401 to the original position. Whenmeasurement of surface plasmon resonance is to be carried out once oneach sample, the measuring unit train 410 is removed from the slideblock 401 there. To the contrast, when measurement of surface plasmonresonance is to be carried out a plurality of times on each sample, theslide block 401 is moved intermittently from left to right again tobring the eight measuring units 10 to the measuring position insequence. Then returning the slide block 401 to the original positionand intermittently moving the slide block 401 from left to right arerepeated according to the number of measurements to be carried out oneach sample.

[0350] The mechanism of linearly moving the slide block 401 describedabove may be also applied to the leaky mode sensor shown in FIG. 9 or 10without limited to the surface plasmon resonance sensor.

[0351] Measuring unit trains shown in FIGS. 26 and 27 may be employed inplace of the measuring unit train 410. The measuring unit train 440shown in FIG. 26 comprises a dielectric bar 441 in the form of arectangular column formed of the same material as the dielectric block11 shown in FIG. 23. A plurality of holes 442 having a closed bottom areformed in the dielectric bar 441, and a metal film layer 12 and asensing medium 14 are formed on the bottom of each hole 442. That is, inthis measuring unit train 440, each hole forms a measuring unit.

[0352] The measuring unit train 440 with this arrangement isadvantageous over that shown in FIG. 23 which is formed by forming aplurality of measuring units 10 one by one and connecting the measuringunits 10 in that the measuring unit 440 can be produced more easily andat a lower cost as compared with that shown in FIG. 23.

[0353] The measuring unit train 450 shown in FIG. 27 comprises aplurality of measuring units 460 fitted in a plurality of unit supportholes 452 formed in a unit support plate 451. Though being basically thesame as the measuring unit 10 shown in FIG. 23, the measuring unit 460is substantially a quadrangular pyramid in shape and accordingly surelyfitted in the unit support hole 452 which is tapered downward.

[0354] In the case of such a measuring unit train 450, a plurality ofmeasuring unit trains 450 may be set on a plate such as a plate 420shown in FIG. 24. Otherwise, one unit support plate 451 loaded with aplurality of measuring units 460 may be fixed on a slide block 401 shownin FIG. 22 and the measuring units 460 may be replaced by another seriesof measuring units 460 after measurement.

[0355] The measuring unit train 410 shown in FIG. 23 or the measuringunit train 440 where the measuring units are fixed to each other not tobe separable is advantageous over the measuring unit train 450 of thisembodiment in that the measuring units can be more easily located inplace and easier to handle since a small measuring unit need not behandled, which contributes to increase in measuring efficiency.

[0356] A surface plasmon resonance sensor in accordance with afourteenth embodiment of the present invention will be described,hereinbelow. FIG. 28 is a side view showing a surface plasmon resonancesensor in accordance with the fourteenth embodiment of the presentinvention. In FIG. 28, elements analogous to those shown in FIG. 2 aregiven the same reference numerals.

[0357] In this embodiment, a support bar 500 which linearly extends inthe direction of arrow Y is employed as the support for supporting aplurality of measuring units 10. A plurality of unit support holes 501are provided at predetermined intervals in the longitudinal direction ofthe support 501, and one measuring unit 502 is held in each unit supportholes 501. Though basically the same as the measuring unit shown in FIG.2, the measuring unit 502 is substantially a quadrangular pyramid inshape and accordingly surely fitted in the unit support hole 501 whichis tapered downward.

[0358] A guide rail 520 is provided below the support bar 500 to extendin parallel to the support bar 500. A slide block 521 is mounted on theguide rail 520 to be slidable along the guide rail 522 in the directionof arrow Y. The slide block 521 is driven back and forth along the guiderail 520 by a drive means 531 mounted thereon.

[0359] A guide rail 522 is fixed on the slide block 521 to extend in thedirection of arrow X in perpendicular to the direction of arrow Y. Aslide block 523 is mounted on the guide rail 522 to be slidable alongthe guide rail 523 in the direction of arrow X. The slide block 523 isdriven back and forth along the guide rail 522 by a drive means 533mounted thereon.

[0360] An optical unit 524 is fixedly mounted on the slide block 523.The optical unit 524 comprises a laser 31 which emits a measuring lightbeam 30, a collimator lens 525 which collimates to a parallel light beamthe light beam 30 which emitted from the laser 31 as a divergent lightbeam, a mirror 526 which reflects the collimated laser beam 30 towardthe interface between the dielectric block 11 and the metal film layer22 of the measuring unit 502, a condenser lens 527 which condenses thelight beam 30 reflected by the mirror 526, a mirror 528 which reflectsthe light beam 30 reflected at the interface, and a photodetector 40which detects the light beam 30 reflected by the mirror 528.

[0361] When surface plasmon resonance is to be measured by the apparatusof this embodiment, the optical unit 524 is held in the standby positionshown by the chained line in FIG. 28. In the standby position, theoptical unit 524 is held in alignment with the first measuring unit 502(the rightmost measuring unit as seen in FIG. 28) in the direction of Yand held out of alignment with the first measuring unit 502 (heldforward or rearward of the first measuring unit 502) in the direction X.

[0362] With the optical unit 524 held in the standby position, aplurality of measuring units 502 are supplied to the unit support holes501 of the support bar 500 by a supply means (not shown), and a sampleliquid 15 is supplied to each measuring unit 502. Then the drive means533 is driven in the regular direction to move the slide block 523 by apredetermined distance in the direction X so that the optical unit 524is brought to a position in the direction X where the optical unit 524can cause the light beam 30 to impinge upon the interface between thedielectric block 11 and the metal film layer 22 of the first measuringunit 502.

[0363] Then the laser 31 is operated to project the light beam 30 to theinterface and the light beam 30 reflected in total internal reflectionat the interface is detected by the photodetector 40, whereby surfaceplasmon resonance is measured. The measurement of surface plasmonresonance is carried out in the same manner as described above.

[0364] After measurement on the first measuring unit 502, the drivemeans 533 is reversed to reverse the slide block 523 by thepredetermined distance in the direction of X so that the optical unit524 is returned to the standby position. Thereafter, the drive means 531is driven to move the slide block 521 leftward by a distance equal topitch of the unit support holes 501. Then the drive means 533 is drivenin the regular direction to move the slide block 523 by thepredetermined distance in the direction X so that the optical unit 524is brought to a position in the direction X where the optical unit 524can cause the light beam 30 to impinge upon the interface between thedielectric block 11 and the metal film layer 22 of the second measuringunit 502.

[0365] Then the laser 31 is operated to project the light beam 30 to theinterface and the light beam 30 reflected in total internal reflectionat the interface is detected by the photodetector 40, whereby surfaceplasmon resonance is measured.

[0366] The same procedure is repeated for the third, fourth, fifth . . .measuring units 502. After measurement on all the measuring units 502 isfinished, the drive means 531 is reversed and the slide block 521 isreturned to the standby position.

[0367] Then by repeating the same procedure, the sample liquid 15 in onemeasuring unit 502 can be subjected to measurement a plurality of times.After a desired number of times of measurement on each sample 15 isfinished, the optical unit 524 is returned to the standby position andthe measuring units 502 is removed from the support bar 500 by adischarge means not shown.

[0368] Then another set of measuring units 502 are supplied to the unitsupport holes 501 of the support bar 500 and another set of sampleliquids 15 are supplied to the measuring units 502. Thereafter, thesample liquids 15 are subjected to measurement in the same manner.

[0369] The mechanism in which the optical unit 524 is moved with themeasuring units kept stationary may be also applied to the leaky modesensor shown in FIG. 9 or 10 without limited to the surface plasmonresonance sensor. Further, it is possible to support a plurality ofmeasuring units along the periphery of a circular support and to causethe measuring optical system to move along the measuring units.

[0370] A surface plasmon resonance sensor in accordance with a fifteenthembodiment of the present invention, where mistaking the result ofmeasurement for one sample for that of a different sample is prevented,will be described, hereinbelow.

[0371]FIG. 29 is a side view showing a surface plasmon resonance sensorin accordance with the fifteenth embodiment of the present invention. Inthe surface plasmon resonance sensor of this embodiment, surface plasmonresonance is measured by the use of a turntable 20 and measuring units80 which are basically the same as those shown in FIG. 21. The samplesare stored in wells of a well-plate 242 similar to that shown in FIG.18, and are dispensed to the measuring units 80 by an automatic samplesupply mechanism 70 provided with dispensing pipettes 71.

[0372] The apparatus of this embodiment is used, for instance, in randomscreening and a sensing medium is fixed on a metal film layer formed onone surface of a dielectric block of the measuring unit 80. Measurementby the apparatus is applied to sample liquids which can contain aspecific material (sample material) which bonds with the sensing medium.Each of the measuring units 80 is provided with an ID barcode 80 brepresenting the production number (a serial number) of the measuringunit 80. The sensing medium will be referred to as a “receptor” and thesample material will be referred to as a “regand”, hereinbelow.

[0373] The surface plasmon resonance sensor of this embodiment isfurther provided with a computer system 600 which manages information onsurface plasmon resonance measurement and a computer system 610 whichoutputs result of measurement on the basis of the output signal S of thephotodetector 40. The computer system 600 may be, for instance, ageneral purpose personal computer comprising a computer body 601, adisplay 602, and a keyboard 603. A first barcode reader 604 which readsthe barcodes 80 b on the measuring units 80 is connected to the computerbody 601. The first barcode reader 604 is placed where a receptor isfixed to the measuring unit 80.

[0374] Also the computer system 610 may be, for instance, a generalpurpose personal computer comprising a computer body 611, a display 612,and a keyboard 613. A second barcode reader 614 which reads the barcodes80 b on the measuring units 80 is connected to the computer body 611.The second barcode reader 614 is placed where a regand is dispensed tothe measuring unit 80.

[0375] A barcode 242 b representing the kind of regands stored in eachwell of the well-plate 242 is applied to the well-plate 242. A thirdbarcode reader 620 which reads the barcode 242 b is connected to thecomputer body 611 of the computer system 610.

[0376] Operation of the surface plasmon resonance sensor of thisembodiment will be described with reference to also FIG. 30,hereinbelow.

[0377] In FIG. 30, the flow of image processing by the computer systems600 and 610 is shown on the right side of FIG. 30, and the flow ofoperation related to the image processing is shown on the left side ofFIG. 30.

[0378] First a measuring unit 80 is produced by a maker. At this time, abarcode 80 b representing the serial number of the measuring unit 80 isapplied to the measuring unit 80. (step P1) At the same time, the serialnumber represented by the barcode 80 b is stored in a predeterminedmemory of the computer system 600, for instance, through the keyboard603 for production control of the measuring units 80. (step Q1) Afterthe barcode 80 b is applied to the measuring unit 80, a metal film layeris formed on the dielectric block of the measuring unit 80. (step P2)

[0379] Then the receptor is fixed on the measuring unit 80. (step P3) Atthis time, the first barcode reader 604 reads the serial numberrepresented by the barcode 80 b of the measuring unit 80 and informationF on the kind of the receptor fixed on the measuring unit 80 is inputthrough the keyboard 603. The information F and the serial number of themeasuring unit 80 are stored and registered in the memory means in thecomputer system 600 correlated to each other. (step Q2) The computersystem 600 transfers the information F to the computer system 610through the internet or an executive communication circuit. The computersystem 610 stores the information F in a predetermined memory means.

[0380] The measuring unit 80 fixed with the receptor is placed on theturntable 20 of the apparatus shown in FIG. 29. The serial numberrepresented by the barcode 80 b on the measuring unit 80 is read by thesecond barcode reader 614 (before or after the measuring unit 80 isplaced on the turntable 20) and input into the computer system 610.(step P4) The computer system 610 reads out the kind of receptorcorresponding to the input serial number on the basis of the informationF stored therein and causes the display 612 to display the kind ofreceptor. The operator can ascertain whether the receptor on themeasuring unit 80 is corrected. (step Q3)

[0381] Then the regands are dispensed to the measuring units 80supported on the turntable 20 and the barcode 242 b representing thekinds of the regands in the wells of the well-plate 242 is read by thethird barcode reader 620. (step P5)

[0382] The regands in the wells of the well-plate 242 are dispensed tothe measuring units 80 in a predetermined order, and at the same time,the measuring units 80 are sent to the regard dispensing position afterpassing through the second barcode reader 614. The computer system 610determines the kind of the regand dispensed to each measuring unit 80 onthe basis of information G from the second barcode reader 614 andinformation H from the third barcode reader 620 and stores and registersthe kind of the regand in correlation with the serial number of themeasuring unit 80. (step Q4)

[0383] After lapse of a predetermined time required for the receptor andthe regand to react with each other, surface plasmon resonancemeasurement is carried out on the measuring units 80 supported on theturntable 20. (step P6) Measurement in this embodiment is carried out inorder to find a regand which can bond to the receptor, and is carriedout on the basis of the output signal S of the photodetector 40 as inthe ninth embodiment shown in FIGS. 14 and 15.

[0384] The computer system 610 determines the result of reaction betweenthe receptors and the regands and determines the bonding state of thereceptors and regands on the basis of the result of reaction. The resultof measurement is stored and registered in the memory means incorrelation to the serial numbers of the measuring units 80. (step Q5)

[0385] The result of measurement thus obtained is displayed by thedisplay 612 of the computer system 610, for instance, as shown in thefollowing table 1. (step Q6) TABLE 1 cup No. receptor regand result ofreaction judgment 22668000 A 10001 5 X 22668001 A 10002 2 X 22668002 A10003 0 X 22668003 A 10004 250 ◯ 22668004 B 10005 10 X 22668005 B 100061340 ◯ . . . . . . . . . . . . . . .

[0386] As described above, in this embodiment, since each of the kind ofthe receptor, the kind of the regand and the result of measurement isstored in correlation with the serial number of the measuring unit 80,there is no fear that measurement is carried out on an unintendedcombination of the receptor and the regand or result of measurement forone combination of the receptor and the regand is mistaken for that of adifferent combination.

[0387] As the identification mark for identifying the measuring units 80from each other, other magnetic recording layers, a semiconductormemory, printed characters read by an OCR or the like may be used.

[0388] Further, though in the embodiment described above, the result ofmeasurement is displayed by a display 612 of the computer system 610,the result of measurement may be printed out on a recording paper.

[0389] A sixteenth embodiment of the present invention will be describedhereinbelow. FIG. 31 is a fragmentary perspective view showing animportant part of a surface plasmon resonance sensor in accordance withthe sixteenth embodiment of the present invention. The surface plasmonresonance sensor of this embodiment differs from that shown in FIG. 5 inthe structure of the optical system which converges the light beam 30and the dielectric block. That is, in this embodiment, a dielectricblock 780 which is like a rectangular prism in shape is employed and themetal film layer 12 is formed on the upper surface of the dielectricblock 780. The optical system comprises a collimator lens 781 whichcollimates the light beam 30 emitted from a laser 31 such as asemiconductor laser as a divergent light beam into a parallel lightbeam, a cylindrical lens 782 which converges the collimated light beam30 only in one direction into a wedge-shaped light beam, and acylindrical lens 783 which converges into a parallel light beam thelight beam 30 which is reflected in total internal reflection at theinterface 780 a between the dielectric block 780 and the metal filmlayer 12 and is divergent only in one direction.

[0390] The cylindrical lens 782 is disposed so that the wedge-shapedlight beam 30 is not focused on the interface 780 a. Accordingly, thespot size of the light beam 30 on the interface 780 a is relativelylarge and about 500 μm×1000 μm (substantially rectangular). Accordingly,the measured value reflects the average of the fine irregularities onthe surface of the metal film layer 12, whereby fluctuation of themeasured values can be suppressed.

[0391] It is possible to fix a sensing medium similar to the sensingmedium 14 shown in FIG. 5 on the surface of the metal film layer 12 andcause the sensing medium to react with a specific material in the sampleliquid 15. Also in this case, when the spot size of the light beam 30 onthe interface 780 a is relatively large, the measured value reflects theaverage of the bonding reaction properties, whereby fluctuation of themeasured values can be suppressed.

[0392] Further, by the use of the cylindrical lens optical system, anoptical system for correcting distortion of the dark line on the lightreceiving face of the photodetector 40 becomes unnecessary, whichreduces the cost of the optical system.

[0393] Though, in the above embodiments, the light beam is caused toimpinge upon the interface between the dielectric block and the metalfilm layer in a defocused state by an arrangement of the optical system,other arrangements may be employed to cause the light beam to impingeupon the interface between the dielectric block and the metal film layerin a defocused state.

[0394] For example, the measuring unit 800 shown in FIG. 32 is arrangedso that the transparent dielectric block 811 forms apart of an opticalsystem which causes the light beam to impinge upon the interface betweenthe dielectric block and the metal film layer in a defocused state. Thatis, the measuring unit 800 comprises a rectangular transparentdielectric block 811, a metal film layer 12 of gold, silver, copper,aluminum or the like formed on the upper surface of the dielectric block811, a sensing medium 14 fixed on the metal film layer 12, and a sampleholder frame 13 which defines a space with closed sides above the metalfilm layer 12.

[0395] When the light beam 30 is caused to enter the rectangulardielectric block 811 as a convergent light beam, astigmatism is causeddue to the shape of the transparent dielectric block 811 and the lightbeam 30 impinges upon the interface 811 a between the dielectric block811 and the metal film layer 12 in a defocused state.

[0396] A seventeenth embodiment of the present invention will bedescribed with reference to FIG. 33, hereinbelow. The sensor utilizingthe phenomenon of attenuation in total internal reflection of theseventeenth embodiment is a leaky mode sensor described above, and alsoin this embodiment, measuring units 990 integrated into a measuring chipare employed. The measuring unit 990 comprises a dielectric block 11 anda clad layer 991 is formed on a surface (the upper surface as seen inFIG. 33) of the dielectric block 11. Further an optical waveguide layer992 is formed on the clad layer 991.

[0397] The dielectric block 11 is formed of, for instance, syntheticresin or optical glass such as BK7. The clad layer 991 is formed ofdielectric material lower than the dielectric block 11 in refractiveindex or metal such as gold. The optical wave guide layer 992 is filmformed of dielectric material such as PMMA which is higher than the cladlayer 991 in refractive index. The thickness of the clad layer 991 is36.5 nm when it is formed of gold film, and the thickness of the opticalwaveguide layer 992 is about 700 nm when it is formed of PMMA.

[0398] In the leaky mode sensor, when the light beam 30 emitted from thelaser 31 is caused to impinge upon the clad layer 991 at an angle notsmaller than an angle of total internal reflection through thedielectric block 11, the light beam 30 is reflected in total internalreflection at the interface 11 a of the dielectric block 11 and the cladlayer 991, and only light having a particular wave number and impingingupon the waveguide layer 992 at a particular angle of incidence comes topropagate through the optical waveguide layer 992 in a waveguide modeafter passing through the clad layer 991. When the waveguide mode isthus excited, almost all the incident light is taken in the opticalwaveguide layer 992 and accordingly, the intensity of light reflected intotal internal reflection at the interface 11 a of the dielectric block11 and the clad layer 991 sharply drops. That is, attenuation in totalinternal reflection occurs.

[0399] Since the wave number of light to be propagated through theoptical waveguide layer 992 in a waveguide mode depends upon therefractive index of the sample 15 on the optical waveguide layer 992,the refractive index and/or the properties of the sample related to therefractive index can be detected on the basis of the angle of incidenceat which the attenuation in total internal reflection occurs. The signalprocessing section 61 quantitatively analyzes the specific material inthe sample 15 under this principle and the result of analysis isdisplayed on a display (not shown).

[0400] The condenser lens 32 is disposed so that the conical light beam30 is not focused on the interface 11 a. Accordingly, the spot size ofthe light beam 30 on the interface 11 a is relatively large and about500 μm×500 μm. Accordingly, the measured value reflects the average ofthe fine irregularities on the surface of the clad layer 991 and/or theoptical waveguide layer 992, whereby fluctuation of the measured valuescan be suppressed.

[0401] A method of measuring surface plasmon resonance in accordancewith an embodiment of the present invention will be described,hereinbelow. FIGS. 34A to 34G are views for illustrating the flow ofcontrol in the surface plasmon resonance measuring method in accordancewith the embodiment of the present invention.

[0402] In FIGS. 34A to 34G, reference numerals 100 and 200 respectivelydenotes a measuring unit and a turntable. The measuring unit 100 isbasically the same as the measuring unit 10 shown in FIG. 1 andcomprises a dielectric block, a metal film layer and a sample holderframe integrated into a measuring chip. Here the measuring unit will bereferred to as “the measuring chip”. The turntable 200 is provided withninety-six (96) chip supporting portions 300 arranged at regular angularintervals (FIGS. 34A to 34G show only part of the ninety-six chipsupporting portions 300), and is intermittently rotated by apredetermined angle equal to the angular intervals of the chipsupporting portions 400 each time. The time intervals betweenintermittent rotations are two seconds in this particular embodiment.

[0403] Positions indicated at F1, F2, F3, F4 and F5 in FIGS. 34A to 34Gare measuring chip supply position, a sensing medium supply position, ameasuring position, a sample injection position and a measuring chipdischarge position, respectively. The sensing material is a materialwhich bonds to a specific material in the sample as the sensing medium14 shown in FIG. 2 and specific examples of the sensing material are asdescribed above.

[0404] As shown in FIG. 34A, a measuring chip 100 in which a sensingmedium fixing film has been fixed on the metal film layer is supplied toa chip supporting portion 300 of the turntable 200 opposed to themeasuring chip supply position F1.

[0405] Then the turntable 200 is once rotated by the predetermined angleso that the measuring chip 100 on the chip supporting portion 300 isbrought to the sensing medium supply position, and a sensing medium issupplied to the measuring chip 100.

[0406] Supply of the sensing medium is carried out in parallel to supplyof the measuring chip 100. When the turntable 200 is intermittentlyrotated 96 times, i.e., when the turntable 200 makes one rotation, theturntable 200 returns to the initial position with all the chipsupporting portions 300 occupied with measuring chips 100 with a sensingmedium. The time required for the turntable 200 to return to the initialposition is 192 seconds (2×96). FIG. 34C shows the turntable 200 in thisstate. Then the surface plasmon resonance sensor is kept in this statefor a predetermined time interval as required.

[0407] Then surface plasmon resonance is measured in the measuringposition F3 on each other measuring chips 100 before supplied with asample, and the output signal S of the photodetector 40 (See, e.g., FIG.2) in this state is detected as a base line of measurement.

[0408] When the turntable 200 is intermittently rotated another time,the measuring chip 100 whose base line is just detected in the measuringposition F3 is brought to the sample injection position F4 shown in FIG.34E and a sample is injected into the measuring chip 100.

[0409] Injection of the sample is carried out in parallel to detectionof the base line. By the time the turntable 200 makes another rotationand returns to the initial position, base line detection and sampleinjection have been carried out on all the measuring chips 100.

[0410] Then as shown in FIG. 34F, surface plasmon resonance measurementis carried out on the measuring chip 100 which is stopped in themeasuring position F3. At this time, the measuring chip 100 is loadedwith a sample and the total reflection attenuation angle θsp isdetected. By the time the turntable 200 is intermittently rotated 96times and returns to the initial position again, surface plasmonresonance measurement has been carried out once on all the measuringchips 100.

[0411] By carrying out surface plasmon resonance measurement on themeasuring chip 100 which is stopped in the measuring position F3 eachtime the turntable 200 is intermittently rotated, measurement can becarried out a plurality of times according to the number of rotations ofthe turntable 200 on each measuring chip 100.

[0412] In accordance with the method of surface plasmon resonancemeasurement of this embodiment, since surface plasmon resonancemeasurement can be carried out on the other measuring chips 100 betweenthe time surface plasmon resonance measurement is carried out onemeasuring chip 100 and the time surface plasmon resonance measurement iscarried out again on the same measuring chip 100, lots of samples can beefficiently measured in a short time.

[0413] When the turntable 200 starts making an n-th rotation, n-thmeasurement is carried out on each measuring chip 100. Each measuringchip 100 is removed from the turntable 200 in the measuring chipdischarge position F5 shown in FIG. 34G after the n-th measurement.

[0414] After injection of the sample into all the measuring chips 100 onthe turntable 200 is finished, the turntable 200 may be kept stopped fora predetermined time to wait for start of reaction between the sampleand the sensing medium.

[0415]FIG. 35 shows an example of result of multiple-time measurement.In FIG. 35, “sampling time” on the abscissa represents the time up toeach measurement after injection of the sample, whereas “amount ofbonded material” on the ordinate represents the amount of the specificmaterial specifically bonded to the sensing medium, which corresponds tochange of surface plasmon resonance detecting signal or movement of theaforesaid dark line.

[0416] As can be understood from FIG. 35, surface plasmon resonance ismeasured a plurality of times, the time required for the amount ofbonded material to be saturated or the like can be known as well as thefinal amount of bonded material.

[0417] Though a method of surface plasmon resonance measurement in whicha turntable 200 is employed has been described, the present inventioncan be applied also to a method where a support is linearly moved backand forth.

1. A measuring apparatus utilizing the phenomenon of attenuation intotal internal reflection comprising a plurality of measuring units eachcomprising a dielectric block, a film layer which is formed on a surfaceof the dielectric block and a sample holder which holds a sample on thesurface of the film layer, a support which supports the measuring units,a light source which emits a light beam, an optical system which causesthe light beam to enter the dielectric block at various angles ofincidence so that the total internal reflection condition is satisfiedat the interface of the dielectric block and the film layer, aphotodetector which detects attenuation in total internal reflection bydetecting the intensity of the light beam which is reflected in totalinternal reflection at the interface, and a drive means which moves thesupport relatively to the optical system and the photodetector to bringthe measuring units in sequence to a predetermined position with respectto the optical system and the photodetector where the total internalreflection condition is satisfied at the interface of the dielectricblock and the film layer and various angles of incidence to theinterface can be obtained.
 2. A measuring apparatus utilizing thephenomenon of attenuation in total internal reflection comprising aplurality of measuring units each comprising a dielectric block, a filmlayer which is formed on a surface of the dielectric block, sensingmaterial which interacts a specific component in a sample and isdisposed on the surface of the film layer, and a sample holder whichholds the sample on the surface of the sensing material, a support whichsupports the measuring units, a light source which emits a light beam,an optical system which causes the light beam to enter the dielectricblock at various angles of incidence so that the total internalreflection condition is satisfied at the interface of the dielectricblock and the film layer, a photodetector which detects attenuation intotal internal reflection by detecting the intensity of the light beamwhich is reflected in total internal reflection at the interface, and adrive means which moves the support relatively to the optical system andthe photodetector to bring the measuring units in sequence to apredetermined position with respect to the optical system and thephotodetector where the total internal reflection condition is satisfiedat the interface of the dielectric block and the film layer and variousangles of incidence to the interface can be obtained.
 3. A measuringapparatus utilizing the phenomenon of attenuation in total internalreflection comprising a plurality of measuring units each comprising adielectric block, a metal film layer which is formed on a surface of thedielectric block and a sample holder which holds a sample on the surfaceof the metal film layer, a support which supports the measuring units, alight source which emits a light beam, an optical system which causesthe light beam to enter the dielectric block at various angles ofincidence so that the total internal reflection condition is satisfiedat the interface of the dielectric block and the metal film layer, aphotodetector which detects attenuation in total internal reflection bysurface plasmon resonance by detecting the intensity of the light beamwhich is reflected in total internal reflection at the interface, and adrive means which moves the support relatively to the optical system andthe photodetector to bring the measuring units in sequence to apredetermined position with respect to the optical system and thephotodetector where the total internal reflection condition is satisfiedat the interface of the dielectric block and the metal film layer andvarious angles of incidence to the interface can be obtained.
 4. Ameasuring apparatus utilizing the phenomenon of attenuation in totalinternal reflection comprising a plurality of measuring units eachcomprising a dielectric block, a metal film layer which is formed on asurface of the dielectric block, sensing material which interacts aspecific component in a sample and is disposed on the surface of themetal film layer, and a sample holder which holds the sample on thesurface of the sensing material, a support which supports the measuringunits, a light source which emits a light beam, an optical system whichcauses the light beam to enter the dielectric block at various angles ofincidence so that the total internal reflection condition is satisfiedat the interface of the dielectric block and the metal film layer, aphotodetector which detects attenuation in total internal reflection bysurface plasmon resonance by detecting the intensity of the light beamwhich is reflected in total internal reflection at the interface, and adrive means which moves the support relatively to the optical system andthe photodetector to bring the measuring units in sequence to apredetermined position with respect to the optical system and thephotodetector where the total internal reflection condition is satisfiedat the interface of the dielectric block and the metal film layer andvarious angles of incidence to the interface can be obtained.
 5. Ameasuring apparatus utilizing the phenomenon of attenuation in totalinternal reflection comprising a plurality of measuring units eachcomprising a dielectric block, a film layer consisting of a clad layerformed on a surface of the dielectric block and an optical waveguidelayer formed on the clad layer, and a sample holder which holds a sampleon the surface of the film layer, a support which supports the measuringunits, a light source which emits a light beam, an optical system whichcauses the light beam to enter the dielectric block at various angles ofincidence so that the total internal reflection condition is satisfiedat the interface of the dielectric block and the clad layer, aphotodetector which detects attenuation in total internal reflection byexcitation of waveguide mode at the optical waveguide layer by detectingthe intensity of the light beam which is reflected in total internalreflection at the interface, and a drive means which moves the supportrelatively to the optical system and the photodetector to bring themeasuring units in sequence to a predetermined position with respect tothe optical system and the photodetector where the total internalreflection condition is satisfied at the interface of the dielectricblock and the clad layer and various angles of incidence to theinterface can be obtained.
 6. A measuring apparatus utilizing thephenomenon of attenuation in total internal reflection comprising aplurality of measuring units each comprising a dielectric block, a filmlayer consisting of a clad layer formed on a surface of the dielectricblock and an optical waveguide layer formed on the clad layer, sensingmaterial which interacts a specific component in a sample and isdisposed on the surface of the film layer, and a sample holder whichholds the sample on the surface of the sensing material, a support whichsupports the measuring units, a light source which emits a light beam,an optical system which causes the light beam to enter the dielectricblock at various angles of incidence so that the total internalreflection condition is satisfied at the interface of the dielectricblock and the clad layer, a photodetector which detects attenuation intotal internal reflection by excitation of waveguide mode at the opticalwaveguide layer by detecting the intensity of the light beam which isreflected in total internal reflection at the interface, and a drivemeans which moves the support relatively to the optical system and thephotodetector to bring the measuring units in sequence to apredetermined position with respect to the optical system and thephotodetector where the total internal reflection condition is satisfiedat the interface of the dielectric block and the clad layer and variousangles of incidence to the interface can be obtained.
 7. A measuringapparatus utilizing the phenomenon of attenuation in total internalreflection as defined in any one of claims 1 to 6 in which the drivemeans moves the support with the optical system and the photodetectorkept stationary.
 8. A measuring apparatus utilizing the phenomenon ofattenuation in total internal reflection as defined in claim 7 in whichthe support is a turntable which supports the measuring units about itsaxis of rotation and the drive means intermittently rotates theturntable.
 9. A measuring apparatus utilizing the phenomenon ofattenuation in total internal reflection as defined in claim 7 in whichthe support supports the measuring units in a row and the drive meansintermittently moves the support in the direction of the row of themeasuring units.
 10. A measuring apparatus utilizing the phenomenon ofattenuation in total internal reflection as defined in any one of claims1 to 6 in which the drive means moves the optical system and thephotodetector with the support kept stationary.
 11. A measuringapparatus utilizing the phenomenon of attenuation in total internalreflection as defined in claim 10 in which the support supports themeasuring units in a row and the drive means intermittently moves theoptical system and the photodetector along the measuring units in a row.12. A measuring apparatus utilizing the phenomenon of attenuation intotal internal reflection as defined in claim 10 in which the supportsupports the measuring units in a row and the drive means intermittentlymoves the optical system and the photodetector along the measuring unitsin a row.
 13. A measuring apparatus utilizing the phenomenon ofattenuation in total internal reflection as defined in claim 8 or 11 inwhich the drive means is provided with a roller bearing which supportsits rotating shaft, and the drive means is arranged to rotate therotating shaft in one direction when a set of measuring units on thesupport are measured and to rotate the rotating shaft in the otherdirection to return the rotating shaft to the original position afterthe measurement on the measuring units to wait in the original positionfor measurement on another set of measuring units.
 14. A measuringapparatus utilizing the phenomenon of attenuation in total internalreflection as defined in claim 9 or 12 in which the measuring units areconnected in a row to form a measuring unit train, and the support isarranged to support the measuring unit train.
 15. A measuring apparatusutilizing the phenomenon of attenuation in total internal reflection asdefined in any one of claims 1 to 14 further comprising an automaticsample feeding means which automatically feeds a sample to the sampleholder of each of the measuring units.
 16. A measuring apparatusutilizing the phenomenon of attenuation in total internal reflection asdefined in any one of claims 1 to 15 in which the dielectric block ofeach measuring unit is fixed to the support while the film layer and thesample holder are integrated with each other to form a measuring chip,and the measuring chips are exchangeable with respect to the dielectricblock.
 17. A measuring apparatus utilizing the phenomenon of attenuationin total internal reflection as defined in claim 16 further comprising ameasuring chip cassette in which plurality of measuring chips arecontained, and a chip supply means which takes out the measuring chipsfrom the measuring chip cassette one by one and mounts each measuringchip on the dielectric block.
 18. A measuring apparatus utilizing thephenomenon of attenuation in total internal reflection as defined in anyone of claims 1 to 15 in which the dielectric block, the film layer andthe sample holder of the measuring unit are integrated with each otherto form a measuring chip which is exchangeable with respect to thesupport.
 19. A measuring apparatus utilizing the phenomenon ofattenuation in total internal reflection as defined in claim 18 furthercomprising a measuring chip cassette in which plurality of measuringchips are contained, and a chip supply means which takes out themeasuring chips from the measuring chip cassette one by one and mountseach measuring chip on the support.
 20. A measuring apparatus utilizingthe phenomenon of attenuation in total internal reflection as defined inany one of claims 1 to 19 in which the optical system is arranged tocause the light beam to enter the dielectric block as a convergent lightbeam or a divergent light beam, and the photodetector is arranged todetect a position of a dark line which is generated in the light beamreflected in total internal reflection at the interface due toattenuation in total reflection.
 21. A measuring apparatus utilizing thephenomenon of attenuation in total internal reflection as defined inclaim 20 in which the optical system is arranged to cause the light beamto impinge upon the interface in a defocused state.
 22. A measuringapparatus utilizing the phenomenon of attenuation in total internalreflection as defined in claim 21 in which the beam diameter of thelight beam as measured on the interface in the direction of movement ofthe support is at least ten times the mechanical positioning accuracy ofthe support.
 23. A measuring apparatus utilizing the phenomenon ofattenuation in total internal reflection as defined in any one of claims1 to 22 in which the measuring units are supported on the upper side ofthe support, the light source is arranged to emit the light beamdownward from above the support, the optical system is provided with areflecting member which reflects upward the light beam toward theinterface.
 24. A measuring apparatus utilizing the phenomenon ofattenuation in total internal reflection as defined in any one of claims1 to 23 in which the measuring units are supported on the upper side ofthe support, the optical system is arranged to cause the light beam toimpinge upon the interface from below the interface, the photodetectoris positioned above the support with its light receiving face directeddownward, and a reflecting member which reflects the light beam,reflected in total internal reflection at the interface, upward towardthe photodetector is provided.
 25. A measuring apparatus utilizing thephenomenon of attenuation in total internal reflection as defined in anyone of claims 1 to 24 further comprising a temperature control meanswhich maintains the temperature of the measuring units at apredetermined temperature before and/or after the measuring units aresupported by the support.
 26. A measuring apparatus utilizing thephenomenon of attenuation in total internal reflection as defined in anyone of claims 1 to 25 further comprising a stirrer means which stirs thesample held by the sample holder of the measuring unit supported by thesupport before attenuation in total internal reflection is detected. 27.A measuring apparatus utilizing the phenomenon of attenuation in totalinternal reflection as defined in any one of claims 1 to 26 in which areference liquid supply means which supplies at least one of themeasuring units supported by the support with reference liquid which hasoptical properties related to the optical properties of the sample isprovided, and data on the state of attenuation in total internalreflection for the sample obtained by the photodetector is corrected onthe basis of data on the state of total internal reflection for thereference liquid.
 28. A measuring apparatus utilizing the phenomenon ofattenuation in total internal reflection as defined in claim 27 in whichthe sample comprises a sample material dissolved in solvent, and thereference liquid supply means supplies the solvent as the referenceliquid.
 29. A measuring apparatus utilizing the phenomenon ofattenuation in total internal reflection as defined in any one of claims1 to 27 further comprising an identification mark provided on each ofthe measuring units, a reading means which reads out the identificationmark from each of the measuring units subjected to the measurement, asample information input means which inputs information on the samplesfed to the respective measuring units, a display means which displaysthe result of the measurement, and a control means which is connected tothe display means, the sample information input means and the readingmeans to store the identification mark for each measuring unit and theinformation on the sample fed to the measuring unit correlated to eachother and causes the display means to display the result of themeasurement on the sample held by a measuring unit in correlation withthe identification mark for the measuring unit and the information onthe sample fed to the measuring unit stored correlated to each other.30. A measuring method employing a measuring apparatus utilizing thephenomenon of attenuation in total internal reflection as defined in anyone of claims 1 to 29, the comprising the steps of detecting attenuationin total internal reflection for the sample held in first one of themeasuring units, moving the support relatively to the optical system andthe photodetector to bring a second one of the measuring units to thepredetermined position with respect to the optical system and thephotodetector and detecting attenuation in total internal reflection forthe sample held in the second one of the measuring units, and moving thesupport relatively to the optical system and the photodetector to bringthe first one of the measuring units again to the predetermined positionwith respect to the optical system and the photodetector and detectingagain attenuation in total internal reflection for the sample held inthe first one of the measuring units.
 31. A measuring apparatusutilizing the phenomenon of attenuation in total internal reflectioncomprising a dielectric block, a film layer which is formed on onesurface of the dielectric block and is brought into contact with asample, a light source which emits a light beam, an optical system whichcauses the light beam to enter the dielectric block in convergent lightso that the total internal reflection condition is satisfied at theinterface of the dielectric block and the film layer and various anglesof incidence of the light beam to the interface of the dielectric blockand the film layer can be obtained, and a photodetector which detectsattenuation in total internal reflection by detecting the intensity ofthe light beam which is reflected in total internal reflection at theinterface, wherein the improvement comprises that the optical system isarranged so that the light beam is not focused on the interface.
 32. Ameasuring apparatus utilizing the phenomenon of attenuation in totalinternal reflection comprising a dielectric block, a metal film layerwhich is formed on one surface of the dielectric block and is broughtinto contact with a sample, a light source which emits a light beam, anoptical system which causes the light beam to enter the dielectric blockin convergent light so that the total internal reflection condition issatisfied at the interface of the dielectric block and the metal filmlayer and various angles of incidence of the light beam to the interfaceof the dielectric block and the metal film layer can be obtained, and aphotodetector which detects attenuation in total internal reflection bysurface plasmon resonance by detecting the intensity of the light beamwhich is reflected in total internal reflection at the interface,wherein the improvement comprises that the optical system is arranged sothat the light beam is not focused on the interface.
 33. A measuringapparatus utilizing the phenomenon of attenuation in total internalreflection comprising a dielectric block, a film layer which is formedof a clad layer formed on a surface of the dielectric block and anoptical waveguide layer formed on the clad layer and is brought intocontact with a sample, a light source which emits a light beam, anoptical system which causes the light beam to enter the dielectric blockin convergent light so that the total internal reflection condition issatisfied at the interface of the dielectric block and the clad layerand various angles of incidence of the light beam to the interface ofthe dielectric block and the clad layer can be obtained, and aphotodetector which detects attenuation in total internal reflection byexcitation of waveguide mode at the optical waveguide layer by detectingthe intensity of the light beam which is reflected in total internalreflection at the interface, wherein the improvement comprises that theoptical system is arranged so that the light beam is not focused on theinterface.
 34. A measuring apparatus utilizing the phenomenon ofattenuation in total internal reflection as defined in any one of claims31 to 33 in which the light beam is at least 500 μm in a cross-sectionalsize at least in one direction on the interface.
 35. A measuringapparatus utilizing the phenomenon of attenuation in total internalreflection as defined in any one of claims 31 to 34 in which the opticalsystem focuses the light beam so that the interface is positionedoutside the focal depth of the light beam.
 36. A measuring apparatusutilizing the phenomenon of attenuation in total internal reflection asdefined in any one of claims 31 to 34 in which the optical system isarranged so that the light beam is not focused on the interface due toits aberration.
 37. A measuring apparatus utilizing the phenomenon ofattenuation in total internal reflection as defined in any one of claims31 to 36 in which the light beam is converged in a conical shape by aspherical lens.
 38. A measuring apparatus utilizing the phenomenon ofattenuation in total internal reflection as defined in any one of claims31 to 36 in which the light beam is converged in a wedge-like shape by acylindrical lens.
 39. A measuring apparatus utilizing the phenomenon ofattenuation in total internal reflection as defined in any one of claims31 to 38 in which a sensing medium which makes bonding reaction with aspecific material in the sample is fixed on the film layer.
 40. Ameasuring apparatus utilizing the phenomenon of attenuation in totalinternal reflection as defined in any one of claims 31 to 39 furthercomprising a sample holder for holding the sample on the film layer. 41.A measuring apparatus utilizing the phenomenon of attenuation in totalinternal reflection as defined in any one of claims 31 to 40 furthercomprising a sample introduction mechanism for introducing the sampleonto the film layer.
 42. A measuring method utilizing the phenomenon ofattenuation in total internal reflection comprising the steps ofbringing a sample into contact with a film layer formed on one surfaceof a dielectric block, causing a light beam to enter the dielectricblock in convergent light so that the total internal reflectioncondition is satisfied at the interface of the dielectric block and thefilm layer and various angles of incidence of the light beam to theinterface of the dielectric block and the film layer can be obtained,and detecting attenuation in total internal reflection by detecting theintensity of the light beam which is reflected in total internalreflection at the interface, wherein the improvement comprises the stepof causing the light beam to enter the dielectric block not to befocused on the interface.
 43. A measuring method utilizing thephenomenon of attenuation in total internal reflection comprising thesteps of bringing a sample into contact with a metal film layer formedon one surface of a dielectric block, causing a light beam to enter thedielectric block in convergent light so that the total internalreflection condition is satisfied at the interface of the dielectricblock and the metal film layer and various angles of incidence of thelight beam to the interface of the dielectric block and the metal filmlayer can be obtained, and detecting attenuation in total internalreflection by surface plasmon resonance by detecting the intensity ofthe light beam which is reflected in total internal reflection at theinterface, wherein the improvement comprises the step of causing thelight beam to enter the dielectric block not to be focused on theinterface.
 44. A measuring method utilizing the phenomenon ofattenuation in total internal reflection comprising the steps ofbringing a sample into contact with an optical waveguide layer formed ona clad layer formed on one surface of a dielectric block, causing alight beam to enter the dielectric block in convergent light so that thetotal internal reflection condition is satisfied at the interface of thedielectric block and the clad layer and various angles of incidence ofthe light beam to the interface of the dielectric block and the cladlayer can be obtained, and detecting attenuation in total internalreflection by detecting the intensity of the light beam which isreflected in total internal reflection at the interface, wherein theimprovement comprises the step of causing the light beam to enter thedielectric block not to be focused on the interface.
 45. A measuringmethod utilizing the phenomenon of attenuation in total internalreflection as defined in any one of claims 42 to 44 in which the lightbeam is caused to enter the dielectric block so as to be at least 500 μmin a cross-sectional size at least in one direction on the interface.